Unit 3 - Genetics

Asexual reproduction in two multicellular organisms:

(a) This relatively simple animal, a hydra, reproduces by budding. The bud, a localized mass of mitotically dividing cells, develops into a small hydra, which detaches from the parent (LM). (b) All the trees in this circle of redwoods arose asexually from a single parent tree, whose stump is in the center of the circle.

What happened to the white flowered plants' genetic contribution to the hybrids?

If it were lost, then the F1 plants could produce only purple-flowered offspring in the F2 generation. But when Mendel allowed the F1 plants to self- or cross-pollinate and planted their seeds, the white-flower trait reappeared in the F2 generation.

In correlating the behavior of chromosomes with that of genes, this chapter will extend what you learned in the past two chapters. First, we'll describe evidence from the fruit fly that strongly supported the chromosome theory. (Although this theory made a lot of sense, it still required experimental evidence.)

Next, we'll explore the chromosomal basis for the transmission of genes from parents to offspring, including what happens when two genes are linked on the same chromosome. Finally, we will discuss some important exceptions to the standard mode of inheritance.

Crossing Over and Synapsis During Prophase I

Prophase I of meiosis is a very busy time. The prophase I cell shown in Figure 13.8 is at a point fairly late in prophase I, when pairing of homologous chromosomes, crossing over, and chromosome condensation have already taken place.

Meiosis II

Sister chromatids separate. Haploid cells with unduplicated chromosomes.

Recombination of Linked Genes: Crossing Over Now, let's explain the results of the Drosophila testcross in Figure 15.9. Recall that most of the offspring from the testcross for body color and wing size had parental phenotypes.

That suggested that the two genes were on the same chromosome, since the occurrence of parental types with a frequency greater than 50% indicates that the genes are linked. About 17% of offspring, however, were recombinants.

While Mendel could readily obtain different pea varieties from seed suppliers, Morgan was probably the first person to want different varieties of the fruit fly. He faced the tedious task of carrying out many matings and then microscopically inspecting large numbers of offspring in search of naturally occurring variant individuals. After many months of this, he complained, "Two years' work wasted. I have been breeding those flies for all that time and I've got nothing out of it." Morgan persisted, however, and was finally rewarded with the discovery of a single male fly with white eyes instead of the usual red.

The phenotype for a character most commonly observed in natural populations, such as red eyes in Drosophila, is called the wild type (Figure 15.3). Traits that are alternatives to the wild type, such as white eyes in Drosophila, are called mutant phenotypes because they are due to alleles assumed to have originated as changes, or mutations, in the wild-type allele.

But Ivanowsky clung to the hypothesis that bacteria caused tobacco mosaic disease. Perhaps, he reasoned, the bacteria were small enough to pass through the filter or made a toxin that could do so.

The second possibility was ruled out when the Dutch botanist Martinus Beijerinck carried out a classic series of experiments that showed that the infectious agent in the filtered sap could replicate (Figure 19.2).

Synthesis of an RNA Transcript:

The three stages of transcription, as shown in Figure 17.8 and described next, are initiation, elongation, and termination of the RNA chain. Study Figure 17.8 to familiarize yourself with the stages and the terms used to describe them.

Meosis II

sister chromatids separate. During another round of cell division, the sister chromatids finally separate; four haploid daughter cells result, containing unduplicated chromosomes

The independent assortment of homologous chromosomes in meiosis.

view figure 13.11

The Results of Mendel's F1 Crosses for Seven Characters in Pea Plants:

view table 14.1

Offspring acquire genes from parents by inheriting chromosomes:

Family friends may tell you that you have your mother's nose or your father's eyes. Of course, parents do not, in any literal sense, give their children a nose, eyes, hair, or any other traits. What, then, is actually inherited?

Genetic Recombination and Linkage:

Meiosis and random fertilization generate genetic variation among offspring of sexually reproducing organisms due to independent assortment of chromosomes, crossing over in meiosis I, and the possibility of any sperm fertilizing any egg (see Concept 13.4). Here we'll examine the chromosomal basis of recombination of alleles in relation to the genetic findings of Mendel and Morgan.

In this chapter, you have learned how DNA molecules are arranged in chromosomes and how DNA replication provides the copies of genes that parents pass to offspring. However, it is not enough that genes be copied and transmitted; the information they carry must be used by the cell. In other words, genes must also be expressed.

. In the next chapter, we will examine how the cell expresses the genetic information encoded in DNA.

Overview of meiosis: how meiosis reduces chromosome number. After the chromosomes duplicate in interphase, the diploid cell divides twice, yielding four haploid daughter cells. This overview tracks just one pair of homologous chromosomes, which for the sake of simplicity are drawn in the condensed state throughout.

(1) Interphase: Pair of homologous chromosomes in diploid parent cell. Pair of duplicated homologous chromosomes. Chromosomes duplicate. Diploid cell with duplicated chromosomes. Sister chromatids.

Crossing over and synapsis in prophase I: a closer look 1) After interphase, the chromosomes have been duplicated, and sister chromatids are held together by proteins called cohesins (purple). Each pair of homologs associate along their length. The DNA molecules of two nonsister chromatids are broken at precisely corresponding points. The chromatin of the chromosomes starts to condense. Maternal sister chromatids Paternal sister chromatids Pair of homologous chromosomes: DNA breaks Cohesins Centromere DNA breaks

(2) A zipper-like protein complex, the synaptonemal complex (green), begins to form, attaching one homolog to the other. The chromatin continues to condense. Synaptonemal complex forming (3) The synaptonemal complex is fully formed; the two homologs are said to be in synapsis. During synapsis, the DNA breaks are closed up when each broken end is joined to the corresponding segment of the nonsister chromatid, producing crossovers. Sister chromatids Sister chromatids Synaptonemal complex Crossovers (4) After the synaptonemal complex disassembles, the homologs move slightly apart from each other but remain attached because of sister chromatid cohesion, even though some of the DNA may no longer be attached to its original chromosome. The points of attachment where crossovers have occurred show up as chiasmata. The chromosomes continue to condense as they move toward the metaphase plate. Chiasmata

Given the prevalence and significance of breast cancer, it is not surprising that it was one of the first cancers for which the role of inheritance was investigated. It turns out that for 5-10% of patients with breast cancer, there is evidence of a strong inherited predisposition. Geneticist Mary-Claire King began working on this problem in the mid-1970s. After 16 years of research, she convincingly demonstrated that mutations in one gene—BRCA1—were associated with increased susceptibility to breast cancer, a finding that flew in the face of medical opinion at the time.

(BRCA stands for breast cancer.) Mutations in that gene or a gene called BRCA2 are found in at least half of inherited breast cancers, and tests using DNA sequencing can detect these mutations. A woman who inherits one mutant BRCA1 allele has a 60% probability of developing breast cancer before the age of 50, compared with only a 2% probability for an individual homozygous for the normal allele.

At first glance, incomplete dominance of either allele seems to provide evidence for the blending hypothesis of inheritance, which would predict that the red or white trait could never reappear among offspring of the pink hybrids. In fact, interbreeding F1 hybrids produces F2 offspring with a phenotypic ratio of one red to two pink to one white.

(Because heterozygotes have a separate phenotype, the genotypic and phenotypic ratios for the F2 generation are the same, 1:2:1.) The segregation of the red-flower and white-flower alleles in the gametes produced by the pinkflowered plants confirms that the alleles for flower color are heritable factors that maintain their identity in the hybrids; that is, inheritance is particulate.

In a cell in which DNA synthesis has occurred, all the chromosomes are duplicated, and therefore each consists of two identical sister chromatids, associated closely at the centromere and along the arms. (

(Even though the chromosomes are duplicated, we still say the cell is diploid, or 2n. This because it has only two sets of information regardless of the number of chromatids, which are merely copies of the information in one set.) Figure 13.4 helps clarify the various terms that we use to describe duplicated chromosomes in a diploid cell.

Although two sickle-cell alleles are necessary for an individual to manifest full-blown sickle-cell disease and thus the condition is considered a recessive one, the presence of one sickle-cell allele can affect the phenotype. Thus, at the organismal level, the normal allele is incompletely dominant to the sickle-cell allele (Figure 14.17). At the molecular level, the two alleles are codominant; both normal and abnormal (sickle-cell) hemoglobins are made in heterozygotes (carriers), who are said to have sickle-cell trait.

(Here the word "trait" is used to distinguish this condition from full-blown sickle-cell disease, thus it is used differently from its definition earlier in the chapter—any variant of a phenotypic character.) Heterozygotes are usually healthy but may suffer some symptoms during long periods of reduced blood oxygen.

In April 1953, Watson and Crick surprised the scientific world with a succinct, one-page paper that reported their molecular model for DNA: the double helix, which has since become the symbol of molecular biology. Watson and Crick, along with Maurice Wilkins, were awarded the Nobel Prize in 1962 for this work.

(Sadly, Rosalind Franklin had died at the age of 37 in 1958 and was thus ineligible for the prize.) The beauty of the double helix model was that the structure of DNA suggested the basic mechanism of its replication.

The Chromosomal Basis of Sex: lthough sex has traditionally been described as a pair of binary categories, we are coming to understand that sex classifications may be less distinct. Here, we use the term sex to mean the classification into a group with a shared set of anatomical and physiological traits.

(The term gender, previously used as a synonym of sex, is now more often used to refer to an individual's own cultural experience of identifying as male, female, or otherwise.) In this sense, sex is determined largely by chromosomes.

Only children who inherit two copies of the Tay-Sachs allele (homozygotes) have the disease. Thus, at the organismal level, the Tay-Sachs allele qualifies as recessive. However, the activity level of the lipid-metabolizing enzyme in heterozygotes is intermediate between the activity level in individuals homozygous for the normal allele and the activity level in individuals with Tay-Sachs disease.

(The term normal is used in the genetic sense to refer to the allele coding for the enzyme that functions properly.) The intermediate phenotype observed at the biochemical level is characteristic of incomplete dominance of either allele. Fortunately, the heterozygote condition does not lead to disease symptoms, apparently because half the normal enzyme activity is sufficient to prevent lipid accumulation in the brain.

Recombination of Unlinked Genes: Independent Assortment of Chromosomes: Mendel learned from crosses in which he followed two characters that some offspring have combinations of traits that do not match those of either parent. For example, consider a cross of a dihybrid pea plant with yellow round seeds, heterozygous for both seed color and seed shape (YyRr), with a plant homozygous for both recessive alleles (with green wrinkled seeds, yyrr).

(This cross acts as a testcross because the results will reveal the genotype of the gametes made in the dihybrid YyRr plant.) Let's represent the cross by the following Punnett square: Notice in this Punnett square that one-half of the offspring are expected to inherit a phenotype that matches either of the phenotypes of the P (parental) generation originally crossed to produce the F1 dihybrid (see Figure 15.2). These matching offspring are called parental types. But two nonparental phenotypes are also found among the offspring. Because these offspring have new combinations of seed shape and color, they are called recombinant types, or recombinants for short.

Mutations can change the phenotype of an organism (as you'll learn in Concept 17.5). And if they occur in germ cells, which give rise to gametes, mutations can be passed on from generation to generation. The vast majority of such changes either have no effect or are harmful, but a very small percentage can be beneficial. In either case, mutations are the original source of the variation on which natural selection operates during evolution and are ultimately responsible for the appearance of new species.

(You'll learn more about this process in Unit Four.) The balance between complete fidelity of DNA replication or repair and a low mutation rate has resulted in new proteins that contribute to different phenotypes. Ultimately, over long periods of time, this process leads to new species and thus to the rich diversity of life we see on Earth today.

A deletion occurs when a chromosomal fragment is lost. The affected chromosome is then missing certain genes. The "deleted" fragment may become attached as an extra segment to a sister or nonsister chromatid, producing a duplication of a portion of that chromosome.

. A chromosomal fragment may also reattach to the original chromosome but in the reverse orientation, producing an inversion. A fourth possible result of chromosomal breakage is for the fragment to join a nonhomologous chromosome, a rearrangement called a translocation.

Enhancers and Specific Transcription Factors: As you can see in Figure 18.8, some control elements, named proximal control elements, are located close to the promoter. (Although some biologists consider proximal control elements part of the promoter, in this text we do not.) The more distant distal control elements, groupings of which are called enhancers, may be thousands of nucleotides upstream or downstream of a gene or even within an intron.

. A given gene may have multiple enhancers, each active at a different time or in a different cell type or location in the organism. Each enhancer, however, is generally associated with only that gene and no other.

Viral Genomes: We usually think of genes as being made of double-stranded DNA, but many viruses defy this convention. Their genomes may consist of double-stranded DNA, single-stranded DNA, double-stranded RNA, or single-stranded RNA, depending on the type of virus.

. A virus is called a DNA virus or an RNA virus based on the kind of nucleic acid that makes up its genome. In either case, the genome is usually organized as a single linear or circular molecule of nucleic acid, although the genomes of some viruses consist of multiple molecules of nucleic acid. The smallest viruses known have only three genes in their genome, while the largest have several hundred to 2,000. For comparison, bacterial genomes contain about 200 to a few thousand genes.

The basic mechanics of transcription and translation are similar for bacteria and eukaryotes, but there is an important difference in the flow of genetic information within the cells. Bacteria do not have nuclei. Therefore, nuclear membranes do not separate bacterial DNA and mRNA from ribosomes and the other protein-synthesizing equipment (Figure 17.4a).

. As you'll see later, this lack of compartmentalization allows translation of an mRNA to begin while its transcription is still in progress. By contrast, eukaryotic cells have nuclei. The presence of a nuclear envelope separates transcription from translation in space and time (Figure 17.4b). Transcription occurs in the nucleus, but the mRNA must be transported to the cytoplasm for translation. Before eukaryotic RNA transcripts from proteincoding genes can leave the nucleus, they are modified in various ways to produce the final, functional mRNA. The transcription of a protein-coding eukaryotic gene results in pre-mRNA, and further processing yields the finished mRNA. The initial RNA transcript from any gene, including those specifying RNA that is not translated into protein, is more generally called a primary transcript.

clone

. Because the cells of the offspring arise via mitosis in the parent, the offspring is usually genetically identical to its parent. An individual that reproduces asexually gives rise to a clone, a group of genetically identical individuals. Genetic differences occasionally arise in asexually reproducing organisms as a result of changes in the DNA called mutations, which we will discuss in Concept 17.5.

A number of physical and chemical agents, called mutagens, interact with DNA in ways that cause mutations. In the 1920s, Hermann Muller discovered that X-rays caused genetic changes in fruit flies, and he used X-rays to make Drosophila mutants for his genetic studies.

. But he also recognized an alarming implication of his discovery: X-rays and other forms of high-energy radiation pose hazards to the genetic material of people as well as laboratory organisms. Mutagenic radiation, a physical mutagen, includes ultraviolet (UV) light, which can cause disruptive thymine dimers in DNA (see Figure 16.19).

Inheritance of X-Linked Genes: The fact that males and females inherit a different number of X chromosomes leads to a pattern of inheritance different from that produced by genes located on autosomes.While there are very few Y-linked genes and many help determine sex, the X chromosomes have genes for many characters unrelated to sex. X-linked genes in humans follow the same pattern of inheritance that Morgan observed for the eye color locus he studied in Drosophila (see Figure 15.4).

. Fathers pass X-linked alleles to all of their daughters but to none of their sons. In contrast, mothers can pass X-linked alleles to both sons and daughters, as shown in Figure 15.7 for the inheritance of a mild X-linked disorder, red-green color blindness.

Mendel chose to track only those characters that occurred in two distinct, alternative forms, such as purple or white flower color. He also made sure that he started his experiments with varieties that were true-breeding—that is, over many generations of self-pollination, these plants had produced only the same variety as the parent plant.

. For example, a plant with purple flowers is true-breeding if the seeds produced by self-pollination in successive generations all give rise to plants that also have purple flowers.

Except for small amounts of DNA in mitochondria and chloroplasts, the DNA of a eukaryotic cell is packaged into chromosomes within the nucleus. Every species has a characteristic number of chromosomes.

. For example, humans have 46 chromosomes in their somatic cells—all cells of the body except the gametes and their precursors. Each chromosome consists of a single long DNA molecule, elaborately coiled in association with various proteins. One chromosome includes several hundred to a few thousand genes, each of which is a precise sequence of nucleotides along the DNA molecule.

Nature and Nurture: The Environmental Impact on Phenotype Another departure from simple Mendelian genetics arises when the phenotype for a character depends on environment as well as genotype. A single tree, locked into its inherited genotype, has leaves that vary in size, shape, and greenness, depending on their exposure to wind and sun.

. For humans, nutrition influences height, exercise alters build, sun-tanning darkens the skin, and experience improves performance on intelligence tests. Even identical twins, who are genetic equals, accumulate phenotypic differences as a result of their unique experiences.

In sexual reproduction, two parents give rise to offspring that have unique combinations of genes inherited from the two parents. In contrast to a clone, offspring of sexual reproduction vary genetically from their siblings and both parents: They are variations on a common theme of family resemblance, not exact replicas.

. Genetic variation like that shown in Figure 13.1 is an important consequence of sexual reproduction.

Generally, the phenotypic range is broadest for polygenic characters. Environment contributes to the quantitative nature of these characters, as we have seen in the continuous variation of skin color

. Geneticists refer to such characters as multifactorial, meaning that many factors, both genetic and environmental, collectively influence phenotype.

The old view that the nuclear contents are like a bowl of amorphous chromosomal spaghetti has given way to a new model of a nucleus with a defined architecture and regulated movements of chromatin. Several lines of evidence suggest that genes that are not being expressed are located in the outer edges of the nucleus, while those that are being expressed are found in its interior region. Relocation of particular genes from their chromosomal territories to transcription factories in the interior may be part of the process of readying genes for transcription.

. How long an individual transcription factory may last has not yet been established. In 2014, the National Institutes of Health announced funding for a new "4D Nucleome" program, which aims to investigate the many fascinating questions addressed by this exciting area of current research.

Scientists have been able to exploit the function of Cas9 by introducing a Cas9-guide RNA complex into a cell they wish to alter (Figure 20.14). The guide RNA in the complex is engineered to be complementary to the "target" gene. Cas9 cuts both strands of the target DNA, and the resulting broken ends of DNA trigger a DNA repair system (similar to that shown in Figure 16.19). When there is no undamaged DNA for the enzymes of the repair system to use as a template, as shown at the bottom left of Figure 20.14, the repair enzymes rejoin the ends, sometimes introducing or removing nucleotides.

. If the cut is directed to a coding portion of the gene, the rejoining process often alters the DNA sequence so that the gene no longer works properly. This technique is a highly effective way for researchers to knock out a given gene in order to study what that gene does, and it has already been used in many organisms, including bacteria, fish, mice, insects, human cells, and various crop plants. Researchers have also modified the technique so that CRISPR-Cas9 can be used to repair a gene that has a mutation (see bottom right of Figure 20.14). They introduce a segment from the normal (functional) gene along with the CRISPRCas9 system. After Cas9 cuts the target DNA, repair enzymes can use the normal DNA segment as a template to repair the target DNA at the break point. This approach is used for gene therapy, which will be discussed later in the chapter.

The F1 plants, of genotype YyRr, exhibit both dominant phenotypes, yellow seeds with round shapes, no matter which hypothesis is correct. The key step in the experiment is to see what happens when F1 plants self-pollinate and produce F2 offspring.

. If the hybrids must transmit their alleles in the same combinations in which the alleles were inherited from the P generation, then the F1 hybrids will produce only two classes of gametes: YR and yr. As shown on the left side of Figure 14.8, this "dependent assortment" hypothesis predicts that the phenotypic ratio of the F2 generation will be 3:1, just as in a monohybrid cross:

Seeing these results, Morgan proposed that some process must occasionally break the physical connection between specific alleles of genes on the same chromosome. Later experiments showed that this process, now called crossing over, accounts for the recombination of linked genes.

. In crossing over, which occurs while replicated homologous chromosomes are paired during prophase of meiosis I, a set of proteins orchestrates an exchange of corresponding segments of one maternal and one paternal chromatid (see Figure 13.9). In effect, when a single crossover occurs, end portions of two nonsister chromatids trade places.

The first hint that extranuclear genes exist came from studies by the German scientist Carl Correns on the inheritance of yellow or white patches on the leaves of an otherwise green plant. In 1909, he observed that the coloration of the offspring was determined only by the maternal parent (the source of eggs) and not by the paternal parent (the source of sperm). Subsequent research showed that such coloration patterns, or variegation, are due to mutations in plastid genes that control pigmentation (Figure 15.18).

. In most plants, a zygote receives all its plastids from the cytoplasm of the egg and none from the sperm, which contributes little more than a haploid set of chromosomes. An egg may contain plastids with different alleles for a pigmentation gene. As the zygote develops, plastids containing wild-type or mutant pigmentation genes are distributed randomly to daughter cells. The pattern of leaf coloration exhibited by a plant depends on the ratio of wild-type to mutant plastids in its various tissues.

Proteins often have a modular architecture consisting of discrete structural and functional regions called domains. One domain of an enzyme, for example, might include the active site, while another might allow the enzyme to bind to a cellular membrane.

. In quite a few cases, different exons code for the different domains of a protein (Figure 17.14).

Programming of traits in the form of DNA: The genetic program is written in the language of DNA, the polymer of four different nucleotides you learned about in Concepts 1.1 and 5.5.

. Inherited information is passed on in the form of each gene's specific sequence of DNA nucleotides, much as printed information is communicated in the form of meaningful sequences of letters. In both cases, the language is symbolic. Just as your brain translates the word apple into a mental image of the fruit, cells translate genes into freckles and other features. Most genes program cells to synthesize specific enzymes and other proteins, whose cumulative action produces an organism's inherited traits. The programming of these traits in the form of DNA is one of the unifying themes of biology.

Normal shortening of telomeres may protect organisms from cancer by limiting the number of divisions that somatic cells can undergo. Cells from large tumors often have unusually short telomeres, as we would expect for cells that have undergone many cell divisions. Further shortening would presumably lead to self-destruction of the tumor cells. Telomerase activity is abnormally high in cancerous somatic cells, suggesting that its ability to stabilize telomere length may allow these cancer cells to persist.

. Many cancer cells do seem capable of unlimited cell division, as do immortal strains of cultured cells (see Concept 12.3). For several years, researchers have studied inhibition of telomerase as a possible cancer therapy. While studies that inhibited telomerase in mice with tumors have led to the death of cancer cells, eventually the cells have restored the length of their telomeres by an alternative pathway. This is an area of ongoing research that may eventually yield useful cancer treatments.

Figure 14.5 illustrates these combinations using a Punnett square, a handy diagrammatic device for predicting the allele composition of offspring from a cross between individuals of known genetic makeup.

. Notice that we use a capital letter to symbolize a dominant allele and a lowercase letter for a recessive allele. In our example, P is the purple-flower allele, and p is the white-flower allele; it is often useful as well to be able to refer to the gene itself as the P/p gene.

Viruses may at first seem very different from mutations as a cause of cancer. However, we now know that viruses can interfere with gene regulation in several ways if they integrate their genetic material into the DNA of a cell. Viral integration may donate an oncogene to the cell, disrupt a tumor-suppressor gene, or convert a proto-oncogene to an oncogene.

. Some viruses produce proteins that inactivate p53 and other tumor-suppressor proteins, making the cell more prone to becoming cancerous. Viruses are powerful biological agents; you'll learn more about them in Chapter 19.

Wild-type Neurospora has modest food requirements. It can grow in the laboratory on a simple solution containing minimal nutrients for growth—inorganic salts, glucose, and the vitamin biotin—incorporated into agar, a support medium. From this so-called minimal medium, wild-type mold cells use their metabolic pathways to produce all the other molecules they need for growth, dividing repeatedly and forming visible colonies of genetically identical cells. As shown in Figure 17.2, Beadle and Tatum generated different "nutritional mutants" of Neurospora cells, each of which was unable to synthesize a particular essential nutrient

. Such cells could not grow on minimal medium but could grow on complete medium, which contains all nutrients needed for growth. For Neurospora, the complete medium consists of the minimal medium supplemented with all 20 amino acids and a few other nutrients. Beadle and Tatum hypothesized that in each nutritional mutant, the gene for the enzyme that synthesizes a particular nutrient had been disabled. This approach resulted in a valuable collection of mutant strains of Neurospora, catalogued by their defect in a particular pathway. Two colleagues of theirs, Adrian Srb and Norman Horowitz, used a collection of arginine-requiring mutants to investigate the biochemical pathway for arginine synthesis in Neurospora (Figure 17.3) Srb and Horowitz pinned down each mutant's defect more specifically, using additional tests to distinguish among three classes of arginine-requiring mutants. Mutants in each class required a different set of compounds along the arginine-synthesizing pathway, which has three steps. These results, and those of many similar experiments done by Beadle and Tatum, suggested that each class was blocked at a different step in this pathway because mutants in that class lacked the enzyme that catalyzes the blocked step.

Thus, these consanguineous ("same blood") matings, indicated in pedigrees by double lines, are more likely to produce offspring homozygous for recessive traits—including harmful ones.

. Such effects can be observed in many types of domesticated and zoo animals that have become inbred.

Bacteria have a single type of RNA polymerase that synthesizes not only mRNA but also other types of RNA that function in protein synthesis, such as ribosomal RNA. In contrast, eukaryotes have at least three types of RNA polymerase in their nuclei; the one used for pre-mRNA synthesis is called RNA polymerase II.

. The other RNA polymerases transcribe RNA molecules that are not translated into protein. In the discussion that follows, we start with the features of mRNA synthesis common to both bacteria and eukaryotes and then describe some key differences.

Although mechanisms for conservative or dispersive DNA replication are not easy to devise, these models remained possibilities until they could be ruled out. After two years of preliminary work at the California Institute of Technology in the late 1950s, Matthew Meselson and Franklin Stahl devised a clever experiment that distinguished between the three models, described in Figure 16.11.

. Their results supported the semiconservative model of DNA replication, as predicted by Watson and Crick, and their experiment is widely recognized among biologists as a classic example of elegant design. The basic principle of DNA replication is conceptually simple. However, the actual process involves some complicated biochemical gymnastics, as we will now see

Figure 18.25a shows a pathway in which an intracellular signal leads to the synthesis of a protein that suppresses the cell cycle. In this case, the signal is damage to the cell's DNA, perhaps as the result of exposure to ultraviolet light. Operation of this signaling pathway blocks the cell cycle until the damage has been repaired. Otherwise, the damage might contribute to tumor formation by causing mutations or chromosomal abnormalities

. Thus, the genes for the components of the pathway act as tumor-suppressor genes. The p53 gene, named for the 53,000-dalton molecular weight of its protein product, is a tumor-suppressor gene. The protein it encodes is a specific transcription factor that promotes the synthesis of cell cycle-inhibiting proteins. That is why a mutation that knocks out the p53 gene, like a mutation that leads to a hyperactive Ras protein, can lead to excessive cell growth and cancer (Figure 18.25b).

Experimental evidence in some type of cells supports a model in which two DNA polymerase molecules, one on each template strand, "reel in" the parental DNA and extrude newly made daughter DNA molecules. In this so-called trombone model, the lagging strand is also looped back through the complex (Figure 16.18).

. Whether the complex moves along the DNA or whether the DNA moves through the complex, either anchored or not, are still open, unresolved questions that are under active investigation. It is also possible that the process varies among species.

A Comparison of Mitosis and Meiosis: Figure 13.10 summarizes the key differences between meiosis and mitosis in diploid cells. Basically, meiosis reduces the number of chromosome sets from two (diploid) to one (haploid), whereas mitosis conserves the number of chromosome sets. Therefore, meiosis produces cells that differ genetically from their parent cell and from each other, whereas mitosis produces daughter cells that are genetically identical to their parent cell and to each other.

1. Synapsis and crossing over. During prophase I, duplicated homologs pair up and crossing over occurs, as described previously and in Figure 13.9. Synapsis and crossing over do not occur during prophase of mitosis 2. Alignment of homologous pairs at the metaphase plate. At metaphase I of meiosis, pairs of homologs are positioned at the metaphase plate, rather than individual chromosomes, as in metaphase of mitosis. 3. Separation of homologs. At anaphase I of meiosis, the duplicated chromosomes of each homologous pair move toward opposite poles, but the sister chromatids of each duplicated chromosome remain attached. In anaphase of mitosis, by contrast, sister chromatids separate.

Figure 13.3

A bdelloid rotifer, an animal that reproduces only asexually

Disorders Caused by Structurally Altered Chromosomes: Many deletions in human chromosomes, even in a heterozygous state, cause severe problems. One such syndrome, known as cri du chat ("cry of the cat"), results from a specific deletion in chromosome 5.

A child born with this deletion is severely intellectually disabled, has a small head with unusual facial features, and has a cry that sounds like the mewing of a distressed cat. Such individuals usually die in infancy or early childhood.

Deletions and duplications are especially likely to occur during meiosis. In crossing over, nonsister chromatids sometimes exchange unequal-sized segments of DNA, so that one partner gives up more genes than it receives (see Figure 21.13). The products of such an unequal crossover are one chromosome with a deletion and one chromosome with a duplication.

A diploid embryo that is homozygous for a large deletion (or has a single X chromosome with a large deletion, in a male) is usually missing a number of essential genes, a condition that is typically lethal. Duplications and translocations also tend to be harmful. In reciprocal translocations, in which segments are exchanged between nonhomologous chromosomes, and in inversions, the balance of genes is not abnormal—all genes are present in their normal doses. Nevertheless, translocations and inversions can alter phenotype because a gene's expression can be influenced by its location among neighboring genes, which can have devastating effects.

Combinatorial Control of Gene Activation: In eukaryotes, the precise control of transcription depends largely on the binding of activators to DNA control elements. Considering the great number of genes that must be regulated in a typical animal or plant cell, the number of completely different nucleotide sequences found in control elements is surprisingly small.

A dozen or so short nucleotide sequences appear again and again in the control elements for different genes. On average, each enhancer is composed of about ten control elements, each of which can bind only one or two specific transcription factors. It is the particular combination of control elements in an enhancer associated with a gene, rather than the presence of a single unique control element, that is important in regulating transcription of the gene.

Axis Establishment: As we mentioned earlier, cytoplasmic determinants in the egg are the substances that initially establish the axes of the Drosophila body. These substances are encoded by genes of the mother, fittingly called maternal effect genes.

A gene classified as a maternal effect gene is one that, when mutant in the mother, results in a mutant phenotype in the offspring, regardless of the offspring's own genotype. In fruit fly development, the mRNA or protein products of maternal effect genes are placed in the egg while it is still in the mother's ovary. When the mother has a mutation in such a gene, she makes a defective gene product (or none at all), and her eggs are defective; when these eggs are fertilized, they fail to develop properly

In humans, the anatomical signs of sex begin to emerge when the embryo is about 2 months old. Before then, the rudiments of the gonads are generic—they can develop into either testes or ovaries, depending on whether or not a Y chromosome is present, and depending on what genes are active.

A gene on the Y chromosome—called SRY, for sex-determining region of Y—is required for the development of testes. In the absence of SRY, the gonads develop into ovaries, even in an XY embryo.

Determining Gene Function: Once they identify a gene of interest, how do scientists determine its function?

A gene's sequence can be compared with sequences in other species. If the function of a similar gene in another species is known, one might suspect that the gene product in question performs a comparable task. Data about the location and timing of gene expression may reinforce the suggested function. To obtain stronger evidence, one approach is to disable the gene and then observe the consequences in the cell or organism.

locus

A gene's specific location along the length of a chromosome is called the gene's locus (plural, loci; from the Latin, meaning "place"). Our genetic endowment (our genome) consists of the genes and other DNA that make up the chromosomes we inherited from our parents.

When considering phenotypes, however, it is often useful to start by focusing on genes that code for polypeptides. In this chapter, you have learned in molecular terms how a typical gene is expressed—by transcription into RNA and then translation into a polypeptide that forms a protein of specific structure and function. Proteins, in turn, bring about an organism's observable phenotype.

A given type of cell expresses only a subset of its genes. This is an essential feature in multicellular organisms: You'd be in trouble if the lens cells in your eyes started expressing the genes for hair proteins, which are normally expressed only in hair follicle cells! Gene expression is precisely regulated, which we'll explore in the next chapter, beginning with the simpler case of bacteria and continuing with eukaryotes.

One reason Mendel probably chose to work with peas is that there are many varieties. For example, one variety has purple flowers, while another variety has white flowers.

A heritable feature that varies among individuals, such as flower color, is called a character. Each variant for a character, such as purple or white color for flowers, is called a trait.

Unlike achondroplasia, which is relatively harmless, some dominant alleles cause lethal diseases. Those that do are much less common than recessive alleles that have lethal effects. A lethal recessive allele is only lethal when homozygous; it can be passed from one generation to the next by heterozygous carriers because the carriers themselves have normal phenotypes.

A lethal dominant allele, however, often causes the death of afflicted individuals before they can mature and reproduce, and in this case the allele is not passed on to future generations.

Fertilization and meiosis alternate in sexual life cycles:

A life cycle is the generation-to-generation sequence of stages in the reproductive history of an organism, from conception to production of its own offspring. In this section, we use humans as an example to track the behavior of chromosomes through the sexual life cycle. We begin by considering the chromosome count in human somatic cells and gametes. We will then explore how the behavior of chromosomes relates to the human life cycle and other types of sexual life cycles.

Most people who have recessive disorders are born to parents who are carriers of the disorder but have a normal phenotype, as is the case shown in the Punnett square in Figure 14.16.

A mating between two carriers corresponds to a Mendelian F1 monohybrid cross, so the predicted genotypic ratio for the offspring is 1 AA:2 Aa :1 aa. Thus, each child has a 1 ⁄ 4 chance of inheriting a double dose of the recessive allele; in the case of albinism, such a child will have albinism. From the genotypic ratio, we also can see that out of three offspring with the normal phenotype (one AA plus two Aa), two are predicted to be heterozygous carriers, a 2 ⁄ 3 chance.

Humans and other mammals have two types of sex chromosomes, designated X and Y. The Y chromosome is much smaller than the X chromosome (Figure 15.5).

A person who inherits two X chromosomes, one from each parent, usually develops anatomy we associate with the "female" sex, while "male" properties are associated with the inheritance of one X chromosome and one Y chromosome (Figure 15.6a). Short segments at either end of the Y chromosome are the only regions that are homologous with regions on the X. These homologous regions allow the X and Y chromosomes in males to pair and behave like homologs during meiosis in the testes.

Most methods for cloning pieces of DNA in the laboratory share certain general features. One common approach uses bacteria, most often Escherichia coli. Recall from Figure 16.12 that the E. coli chromosome is a large, circular molecule of DNA. In addition, E. coli and many other bacteria also have plasmids, small, circular DNA molecules that are replicated separately.

A plasmid has only a small number of genes; these genes may be useful when the bacterium is in a particular environment but may not be required for survival or reproduction under most conditions.

If the operator is the operon's switch for controlling transcription, how does this switch work? By itself, the trp operon is turned on; that is, RNA polymerase can bind to the promoter and transcribe the genes of the operon. The trp operon can be switched off by a protein that is called the trp repressor.

A repressor binds to the operator and blocks attachment of RNA polymerase to the promoter, preventing transcription of the genes (Figure 18.3b). A repressor protein is specific for the operator of a particular operon. For example, the trp repressor, which switches off the trp operon by binding to the trp operator, has no effect on other operons in the E. coli genome.

Making Multiple Copies of a Gene or Other DNA Segment: A molecular biologist studying a particular gene or group of genes faces a challenge. Naturally occurring DNA molecules are very long, and a single molecule usually carries hundreds or even thousands of genes. Moreover, in many eukaryotic genomes, protein-coding genes occupy only a small proportion of the chromosomal DNA, the rest being noncoding nucleotide sequences.

A single human gene, for example, might constitute only 1/100,000 of a chromosomal DNA molecule. As a further complication, the distinctions between a gene and the surrounding DNA are subtle, consisting only of differences in nucleotide sequence. To work directly with specific genes, scientists have developed methods for preparing well-defined segments of DNA in multiple identical copies, a process called DNA cloning

mosome in this way, the viral DNA is known as a prophage. One prophage gene codes for a protein that prevents transcription of most of the other prophage genes. Thus, the phage genome is mostly silent within the bacterium. Every time the E. coli cell prepares to divide, it replicates the phage DNA along with its own chromosome such that each daughter cell inherits a prophage.

A single infected cell can quickly give rise to a large population of bacteria carrying the virus in prophage form. This mechanism enables viruses to propagate without killing the host cells on which they depend.

A gene located on either sex chromosome is called a sexlinked gene. The human X chromosome contains approximately 1,100 genes, which are called X-linked genes, while genes located on the Y chromosome are called Y-linked genes. On the human Y chromosome, researchers have identified 78 genes that code for about 25 proteins (some genes are duplicates).

About half of these genes are expressed only in the testis, and some are required for normal testicular functioning and the production of normal sperm. The Y chromosome is passed along virtually intact from a father to all his sons. Because there are so few Y-linked genes, very few disorders are transferred from father to son on the Y chromosome.

In eukaryotes, the subunits are made in the nucleolus. Ribosomal RNA genes are transcribed, and the RNA is processed and assembled with proteins imported from the cytoplasm. Completed ribosomal subunits are then exported via nuclear pores to the cytoplasm. In both bacteria and eukaryotes, a large and a small subunit join to form a functional ribosome only when attached to an mRNA molecule.

About one-third of the mass of a ribosome is made up of proteins; the rest consists of three rRNA molecules (in bacteria) or four (in eukaryotes). Because most cells contain thousands of ribosomes, rRNA is the most abundant type of cellular RNA.

Since we understand the progression of this type of cancer, routine screenings (colonoscopies, for example) are recommended to identify and remove any suspicious polyps. The colorectal cancer mortality rate has been declining for the past 20 years due to increased screening and improved treatments. Treatments for other cancers have improved as well.

Advances in the sequencing of DNA and mRNA allow medical researchers to compare the genes expressed by different types of tumors and by the same type in different people. These comparisons have led to personalized treatments based on the molecular characteristics of a person's tumor.

To figure out the probability that an F2 plant from a monohybrid cross will be heterozygous rather than homozygous, we need to invoke a second rule. Notice in Figure 14.9 that the dominant allele can come from the egg and the recessive allele from the sperm, or vice versa. That is, F1 gametes can combine to produce Rr offspring in two mutually exclusive ways: For any particular heterozygous F2 plant, the dominant allele can come from the egg or the sperm, but not from both.

According to the addition rule, the probability that any one of two or more mutually exclusive events will occur is calculated by adding their individual probabilities. As we have just seen, the multiplication rule gives us the individual probabilities that we will now add together. The probability for one possible way of obtaining an F2 heterozygote—the dominant allele from the egg and the recessive allele from the sperm—is 1 ⁄ 4.

Third, if the two alleles at a locus differ, then one, the dominant allele, determines the organism's appearance; the other, the recessive allele, has no noticeable effect on the organism's appearance.

Accordingly, Mendel's F1 plants had purple flowers because the allele for that trait is dominant and the allele for white flowers is recessive

Eukaryotic genes that are co-expressed, such as genes coding for the enzymes of a metabolic pathway, are typically scattered over different chromosomes. Here, coordinate gene expression depends on every gene of a dispersed group having a specific combination of control elements.

Activator proteins in the nucleus that recognize the control elements bind to them, promoting simultaneous transcription of the genes, no matter where they are in the genome.

Protein Folding and Post-Translational Modifications: During its synthesis, a polypeptide chain begins to coil and fold spontaneously as a consequence of its amino acid sequence (primary structure), forming a protein with a specific shape: a three-dimensional molecule with secondary and tertiary structure (see Figure 5.18). Thus, a gene determines primary structure, which in turn determines shape.

Additional steps—post-translational modifications—may be required before the protein can begin doing its particular job in the cell. Certain amino acids may be chemically modified by the attachment of sugars, lipids, phosphate groups, or other additions. Enzymes may remove one or more amino acids from the leading (amino) end of the polypeptide chain. In some cases, a polypeptide chain may be enzymatically cleaved into two or more pieces. In other cases, two or more polypeptides that are synthesized separately may come together, if the protein has quaternary structure; an example is hemoglobin (see Figure 5.18).

The cases described above involve chromatin remodeling in large regions of the chromosome. Because chromatin structure affects transcription and thus gene expression, RNA-based regulation of chromatin structure is sure to play an important role in gene regulation.

Additionally, some experimental evidence supports the idea of an alternate role for lncRNAs in which they can act as a scaffold, bringing together DNA, proteins, and other RNAs into complexes. These associations may act either to condense chromatin or, in some cases, to help bring the enhancer of a gene together with mediator proteins and the gene's promoter, activating gene expression in a more direct fashion.

An extra X chromosome in a male, producing XXY, occurs approximately once in every 500 to 1,000 live male births. People with this disorder, called Klinefelter syndrome, have male sex organs, but the testes are abnormally small and the man is sterile. Even though the extra X is inactivated, some breast enlargement and other female body characteristics are common.

Affected individuals may have subnormal intelligence. About 1 of every 1,000 males is born with an extra Y chromosome (XYY). These males undergo normal sexual development and do not exhibit any well-defined syndrome, but tend to be taller than average.

A number of human X-linked disorders are much more serious than color blindness, such as Duchenne muscular dystrophy, which affects about one out of 3,500 males born in the United States. The disease is characterized by a progressive weakening of the muscles and loss of coordination.

Affected individuals rarely live past their early 20s. Researchers have traced the disorder to the absence of a key muscle protein called dystrophin and have mapped the gene for this protein to a specific locus on the X chromosome.

The sequence of events leading up to that point is shown in more detail in Figure 13.9.

After interphase, the chromosomes have been duplicated and the sister chromatids are held together by proteins called cohesins. 1) Early in prophase I, the two members of a homologous pair associate loosely along their length. Each gene on one homolog is aligned precisely with the corresponding allele of that gene on the other homolog. The DNA of two nonsister chromatids—one maternal and one paternal—is broken by specific proteins at precisely matching points. 2) Next, the formation of a zipper-like structure called the synaptonemal complex holds one homolog tightly to the other. 3) During this association, called synapsis, the DNA breaks are closed up so that each broken end is joined to the corresponding segment of the nonsister chromatid. Thus, a paternal chromatid is joined to a piece of maternal chromatid beyond the crossover point, and vice versa. 4) These points of crossing over become visible as chiasmata (singular, chiasma) after the synaptonemal complex disassembles and the homologs move slightly apart from each other. The homologs remain attached because sister chromatids are still held together by sister chromatid cohesion, even though some of the DNA may no longer be attached to its original chromosome. At least one crossover per chromosome must occur in order for the homologous pair to stay together as it moves to the metaphase I plate, for reasons that will be explained shortly.

The fact that mitochondrial disorders are inherited only from the mother suggests a way to avoid passing along these disorders. The chromosomes from the egg of an affected mother could be transferred to an egg of a healthy donor that has had its own chromosomes removed. This "two-mother" egg could then be fertilized by a sperm from the prospective father and transplanted into the womb of the prospective mother, becoming an embryo with three parents.

After optimizing conditions for this approach in monkeys, researchers reported in 2013 that they have successfully carried out this procedure on human eggs. More research will be necessary to optimize experimental conditions for the health of the embryo, and eventual use of this procedure would require approval by the relevant federal agencies.

DNA technology and other modern biochemical methods enabled the researchers to test whether the bicoid product, a protein called Bicoid, is in fact a morphogen that determines the anterior end of the fly. The first question they asked was whether the mRNA and protein products of this gene are located in the egg in a position consistent with the hypothesis. They found that bicoid mRNA is highly concentrated at the extreme anterior end of the mature egg (Figure 18.22).

After the egg is fertilized, the mRNA is translated into protein. The Bicoid protein then diffuses from the anterior end toward the posterior, resulting in a gradient of protein within the early embryo, with the highest concentration at the anterior end. These results are consistent with the hypothesis that Bicoid protein specifies the fly's anterior end. To test the hypothesis more specifically, scientists injected pure bicoid mRNA into various regions of early embryos. The protein that resulted from its translation caused anterior structures to form at the injection sites

At each end of a replication bubble is a replication fork, a Y-shaped region where the parental strands of DNA are being unwound. Several kinds of proteins participate in the unwinding (Figure 16.13). Helicases are enzymes that untwist the double helix at the replication forks, separating the two parental strands and making them available as template strands.

After the parental strands separate, single-strand binding proteins bind to the unpaired DNA strands, keeping them from re-pairing. The untwisting of the double helix causes tighter twisting and strain ahead of the replication fork. Topoisomerase is an enzyme that helps relieve this strain by breaking, swiveling, and rejoining DNA strands.

Eukaryotic gene expression is regulated at many stages:

All organisms, whether prokaryotes or eukaryotes, must regulate which genes are expressed at any given time. Both unicellular organisms and the cells of multicellular organisms continually turn genes on and off in response to signals from their external and internal environments. Regulation of gene expression is also essential for cell specialization in multicellular organisms, which are made up of different types of cells. To perform its own distinct role, each cell type must maintain a specific program of gene expression in which certain genes are expressed and others are not.

The Law of Independent Assortment: Mendel derived the law of segregation from experiments in which he followed only a single character, such as flower color.

All the F1 progeny produced in his crosses of true-breeding parents were monohybrids, meaning that they were heterozygous for the one particular character being followed in the cross. We refer to a cross between such heterozygotes as a monohybrid cross

Correlating Behavior of a Gene's Alleles with Behavior of a Chromosome Pair: Morgan mated his white-eyed male fly with a red-eyed female. All the F1 offspring had red eyes, suggesting that the wild-type allele is dominant. When Morgan bred the F1 flies to each other, he observed the classical 3:1 phenotypic ratio among the F2 offspring. However, there was a surprising additional result: The white-eye trait showed up only in males.

All the F2 females had red eyes, while half the males had red eyes and half had white eyes. Therefore, Morgan concluded that somehow a fly's eye color was linked to its sex. (If the eye color gene were unrelated to sex, half of the white-eyed flies would have been female.)

This law applies only to genes (allele pairs) located on different chromosomes (that is, on chromosomes that are not homologous) or, alternatively, to genes that are very far apart on the same chromosome. (This will be explained in Concept 15.3, along with the more complex inheritance patterns of genes located near each other, alleles of which tend to be inherited together.)

All the pea characters Mendel chose for analysis were controlled by genes on different chromosomes or were far apart on the same chromosome; this situation greatly simplified interpretation of his multicharacter pea crosses. All the examples we consider in the rest of this chapter involve genes located on different chromosomes.

Alleles (alternative versions) of a gene: This diagram shows a pair of homologous chromosomes in an F1 hybrid pea plant, with the actual DNA sequence from the flower-color allele of each chromosome. The paternally inherited chromosome (blue) has an allele for purple flowers, which codes for a protein that indirectly controls synthesis of purple pigment. The maternally inherited chromosome (red) has an allele for white flowers, which results in no functional protein being made

Allele for purple flowers Locus for flower-color gene > pair of homologous chromosomes. Allele for white flowers DNA with nucleotide sequence CTAAATCGGT DNA with nucleotide sequence ATAAATCGGT Through a series of steps, this DNA sequence results in production of an enzyme that helps synthesize purple pigment. This DNA sequence results in the absence of the enzyme. One purple-flower allele results in sufficient pigment for purple flowers.

The Relationship Between Dominance and Phenotype: We've now seen that the relative effects of two alleles range from complete dominance of one allele, to incomplete dominance of either allele, to codominance of both alleles. It is important to understand that an allele is called dominant because it is seen in the phenotype, not because it somehow subdues a recessive allele.

Alleles are simply variations in a gene's nucleotide sequence (see Figure 14.4). When a dominant allele coexists with a recessive allele in a heterozygote, they do not actually interact at all. It is in the pathway from genotype to phenotype that dominance and recessiveness come into play.

Cloned organisms and stem cells are useful for basic research and other applications

Along with advances in DNA technology, scientists have been developing and refining methods for cloning whole multicellular organisms from single cells. In this context, cloning produces one or more organisms that are genetically identical to the "parent" that donated the single cell. This is often called organismal cloning to differentiate it from gene cloning and, more significantly, from cell cloning—the division of an asexually reproducing cell such as a bacterium into a group of genetically identical cells. (The common theme is that the product is genetically identical to the parent. In fact, the word clone comes from the Greek klon, meaning "twig.") The current interest in organismal cloning arises primarily from its ability to generate stem cells.

variation

Along with inherited similarity, there is also variation. What are the biological mechanisms leading to the "family resemblance" evident among the family members in the photo? A detailed answer to this question eluded biologists until the advance of genetics in the 20th century.

The many avian flu viruses carried by wild and domestic birds pose a potential long-term threat. A case in point is an H5N1 virus; the first transmission of H5N1 from birds to humans was documented in Hong Kong in 1997. Since then, the overall mortality rate due to H5N1 has been greater than 50% of those infected—an alarming number.

Also, the host range of H5N1 is expanding, which provides increasing chances for reassortment between different strains. If the H5N1 avian flu virus evolves so that it can spread easily from person to person, it could represent a major global health threat akin to that of the 1918 pandemic.

Gene cloning is useful for two basic purposes: to make many copies of, or amplify, a particular gene and to produce a protein product from it (see Figure 20.4). Researchers can isolate copies of a cloned gene from bacteria for use in basic research or to endow another organism with a new metabolic capability, such as pest resistance. For example, a resistance gene present in one crop species might be cloned and transferred into plants of another species. (Such organisms are called genetically modified, or GM for short; they will be discussed later in the chapter.)

Alternatively, a protein with medical uses, such as human growth hormone, can be harvested in large quantities from cultures of bacteria carrying a cloned gene for the protein. (We'll describe the techniques for expressing cloned genes later.) Since one gene is only a very small part of the total DNA in a cell, the ability to amplify such rare DNA fragments is crucial for any application involving a single gene.

Initiation of Translation and mRNA Degradation: Translation is another opportunity for regulating gene expression; it occurs most commonly at the initiation stage (see Figure 17.19). For some mRNAs, the initiation of translation can be blocked by regulatory proteins that bind to specific sequences or structures within the untranslated region (UTR) at the 5′ or 3′ end, preventing the attachment of ribosomes. (Recall from Concept 17.3 that both the 5′ cap and the poly-A tail of an mRNA molecule are important for ribosome binding.)

Alternatively, translation of all the mRNAs in a cell may be regulated simultaneously. In a eukaryotic cell, such "global" control usually involves the activation or inactivation of one or more of the protein factors required to initiate translation. This mechanism plays a role in starting translation of mRNAs that are stored in eggs. Just after fertilization, translation is triggered by the sudden activation of translation initiation factors. The response is a burst of synthesis of the proteins encoded by the stored mRNAs. Some plants and algae store mRNAs during periods of darkness; light then triggers the reactivation of the translational apparatus.

Though interphase chromatin is generally much less condensed than the chromatin of mitotic chromosomes, it shows several of the same levels of higher-order packing. Some of the chromatin comprising a chromosome seems to be present as a 10-nm fiber, but much is compacted into a 30-nm fiber, which in some regions is further folded into looped domains. Early on, biologists assumed that interphase chromatin was a tangled mass in the nucleus, like a bowl of spaghetti, but this is far from the case.

Although an interphase chromosome lacks an obvious scaffold, its looped domains appear to be attached to the nuclear lamina, on the inside of the nuclear envelope, and perhaps also to fibers of the nuclear matrix. These attachments may help organize regions of chromatin where genes are active. The chromatin of each chromosome occupies a specific restricted area within the interphase nucleus, and the chromatin fibers of different chromosomes do not appear to be entangled (Figure 16.23b)

The Watson-Crick model took into account Chargaff's ratios and ultimately explained them. Wherever one strand of a DNA molecule has an A, the partner strand has a T. Similarly, a G in one strand is always paired with a C in the complementary strand. Therefore, in the DNA of any organism, the amount of adenine equals the amount of thymine, and the amount of guanine equals the amount of cytosine. (Modern DNA sequencing techniques have confirmed that the amounts are exactly equal.)

Although the base-pairing rules dictate the combinations of nitrogenous bases that form the "rungs" of the double helix, they do not restrict the sequence of nucleotides along each DNA strand. The linear sequence of the four bases can be varied in countless ways, and each gene has a unique base sequence.

After attending the university, Mendel returned to the monastery and was assigned to teach at a local school, where several other instructors were enthusiastic about scientific research. In addition, his fellow monks shared a long-standing fascination with the breeding of plants. Around 1857, Mendel began breeding garden peas in the abbey garden to study inheritance.

Although the question of heredity had long been a focus of curiosity at the monastery, Mendel's fresh approach allowed him to deduce principles that had remained elusive to others.

Probability laws govern Mendelian inheritance: Mendel's laws of segregation and independent assortment reflect the same rules of probability that apply to tossing coins, rolling dice, and drawing cards from a deck. The probability scale ranges from 0 to 1.

An event that is certain to occur has a probability of 1, while an event that is certain not to occur has a probability of 0. With a coin that has heads on both sides, the probability of tossing heads is 1, and the probability of tossing tails is 0.

What happens if we mate black Labs that are heterozygous for both genes (BbEe)?

Although the two genes affect the same phenotypic character (coat color), they follow the law of independent assortment. Thus, our breeding experiment represents an F1 dihybrid cross, like those that produced a 9:3:3:1 ratio in Mendel's experiments. We can use a Punnett square to represent the genotypes of the F2 offspring (Figure 14.12). As a result of epistasis, the phenotypic ratio among the F2 offspring is 9 black to 3 chocolate to 4 yellow Labs. Other types of epistatic interactions produce different ratios, but all are modified versions of 9:3:3:1.

A chromosome consists of a DNA molecule packed together with proteins: Now that you have learned about the structure and replication of DNA, let's take a step back and examine how DNA is packaged into chromosomes, the structures that carry genetic information. The main component of the genome in most bacteria is one double-stranded, circular DNA molecule that is associated with a small amount of protein.

Although we refer to this structure as the bacterial chromosome, it is very different from a eukaryotic chromosome, which consists of one linear DNA molecule associated with a large amount of protein. In E. coli, the chromosomal DNA consists of about 4.6 million nucleotide pairs, representing about 4,400 genes. This is 100 times more DNA than is found in a typical virus, but only about one-thousandth as much DNA as in a human somatic cell. Still, that is a tremendous amount of DNA to be packaged in such a small container.

The development of female gonads requires a gene called WNT4 (on chromosome 1, an autosome), which encodes a protein that promotes ovary development.

An embryo that is XY but has extra copies of the WNT4 gene can develop rudimentary female gonads. Overall, sex is determined by the interactions of a network of gene products like these.

Since that time, researchers have cloned numerous other mammals, including mice, cats, cows, horses, pigs, dogs, and monkeys. In most cases, their goal has been the production of new individuals; this is known as reproductive cloning. We have already learned a lot from such experiments. For example, cloned animals of the same species do not always look or behave identically. In a herd of cows cloned from the same line of cultured cells, certain cows are dominant in behavior and others are more submissive. Another example of nonidentity in clones is the first cloned cat, named CC for Carbon Copy (Figure 20.18). She has a calico coat, like her single female parent, but the color and pattern are different because of random X chromosome inactivation, which is a normal occurrence during embryonic development (see Figure 15.8).

And identical human twins, which are naturally occurring "clones," are always slightly different. Clearly, environmental influences and random phenomena play a significant role during development.

Aneuploidy of Sex Chromosomes

Aneuploid conditions involving sex chromosomes appear to upset the genetic balance less than those involving autosomes. This may be because the Y chromosome carries relatively few genes. Also, extra copies of the X chromosome simply become inactivated as Barr bodies

In the past 20 years, much effort has gone into developing drugs to treat HIV. Currently, multidrug treatments, sometimes called "cocktails," are considered to be most effective. Such treatments commonly include a combination of two nucleoside mimics and a protease inhibitor, which interferes with an enzyme required for assembly of the viruses.

Another effective treatment involves a drug called maraviroc, which blocks a protein on the surface of human immune cells that helps bind the HIV virus (see Figure 7.8). This drug has also been used successfully to prevent infection in individuals who either have been exposed to, or are at risk of exposure to, HIV

Bacterial Expression Systems: Getting a cloned eukaryotic gene to function in bacterial host cells can be difficult because certain aspects of gene expression are different in eukaryotes and bacteria. To overcome differences in promoters and other DNA control sequences, scientists usually employ an expression vector, a cloning vector that contains a highly active bacterial promoter just upstream of a restriction site where the eukaryotic gene can be inserted in the correct reading frame. The bacterial host cell will recognize the promoter and proceed to express the foreign gene now linked to that promoter. Such expression vectors allow the synthesis of many eukaryotic proteins in bacterial cells.

Another problem with expressing cloned eukaryotic genes in bacteria is the presence of noncoding regions (introns) in most eukaryotic genes (see Concept 17.3). Introns can make a eukaryotic gene very long and unwieldy, and they prevent correct expression of the gene by bacterial cells, which do not have RNA-splicing machinery. This problem can be surmounted by using a form of the gene that includes only the exons. (This is called complementary DNA, or cDNA; see Figure 20.10.)

The number of chromosomes in a single set is represented by n.

Any cell with two chromosome sets is called a diploid cell and has a diploid number of chromosomes, abbreviated 2n. For humans, the diploid number is 46 (2n = 46), the number of chromosomes in our somatic cells.

Mismatched nucleotides sometimes evade proofreading by a DNA polymerase. In mismatch repair, other enzymes remove and replace incorrectly paired nucleotides that have resulted from replication errors. Researchers highlighted the importance of such repair enzymes when they found that a hereditary defect in one of them is associated with a form of colon cancer.

Apparently, this defect allows cancer-causing errors to accumulate in the DNA faster than normal.

Elongation of the RNA Strand:

As RNA polymerase moves along the DNA, it untwists the double helix, exposing about 10-20 DNA nucleotides at a time for pairing with RNA nucleotides (Figure 17.10). The enzyme adds nucleotides to the 3¿ end of the growing RNA molecule as it continues along the double helix. . In the wake of this advancing wave of RNA synthesis, the new RNA molecule peels away from its DNA template, and the DNA double helix re-forms. Transcription progresses at a rate of about 40 nucleotides per second in eukaryotes.

Chromatin undergoes striking changes in its degree of packing during the course of the cell cycle (see Figure 12.7). In interphase cells stained for light microscopy, the chromatin usually appears as a diffuse mass within the nucleus, suggesting that the chromatin is highly extended.

As a cell prepares for mitosis, its chromatin coils and folds up (condenses), eventually forming a characteristic number of short, thick metaphase chromosomes that are distinguishable from each other with the light microscope (Figure 16.23a).

Anticodons are conventionally written 3' -> 5' to align properly with codons written 5' -> 3' (see Figure 17.15). (For base pairing, RNA strands must be antiparallel, like DNA.) As an example of how tRNAs work, consider the mRNA codon 5'-GGC-3', which is translated as the amino acid glycine. The tRNA that base-pairs with this codon by hydrogen bonding has 3'-CCG-5' as its anticodon and carries glycine at its other end (see the incoming tRNA approaching the ribosome in Figure 17.15).

As an mRNA molecule is moved through a ribosome, glycine will be added to the polypeptide chain whenever the codon 5¿-GGC-3¿ is presented for translation. Codon by codon, the genetic message is translated as tRNAs position each amino acid in the order prescribed, and the ribosome adds that amino acid onto the growing polypeptide chain. The tRNA molecule is a translator in the sense that, in the context of the ribosome, it can read a nucleic acid word (the mRNA codon) and interpret it as a protein word (the amino acid)

Regulation of Transcription Initiation: Chromatin-modifying enzymes provide initial control of gene expression by making a region of DNA either more or less able to bind the transcription machinery. Once the chromatin of a gene is optimally modified for expression, the initiation of transcription is the next major step at which gene expression is regulated.

As in bacteria, the regulation of transcription initiation in eukaryotes involves proteins that bind to DNA and either facilitate or inhibit binding of RNA polymerase. The process is more complicated in eukaryotes, however. Before looking at how eukaryotic cells control their transcription, let's review the structure of a eukaryotic gene.

Because of the different effects of dominant and recessive alleles, an organism's traits do not always reveal its genetic composition. Therefore, we distinguish between an organism's appearance or observable traits, called its phenotype, and its genetic makeup, its genotype.

As shown in Figure 14.5 for the case of flower color in pea plants, PP and Pp plants have the same phenotype (purple flowers) but different genotypes. Figure 14.6 reviews these terms. Note that the term phenotype refers to physiological traits as well as traits that relate directly to appearance. For example, one pea variety lacks the normal ability to self-pollinate, which is a phenotypic trait (called non-self-pollination).

Alterations of chromosome number or structure cause some genetic disorders:

As you have learned so far in this chapter, the phenotype of an organism can be affected by small-scale changes involving individual genes. Random mutations are the source of all new alleles, which can lead to new phenotypic traits.

Even morphogenesis, the shaping of the organism, can be traced back to changes in the shape, motility, and other characteristics of the cells that make up various regions of the embryo.

As you have seen, the activities of a cell depend on the genes it expresses and the proteins it produces. Almost all cells in an organism have the same genome; therefore, differential gene expression results from the genes being regulated differently in each cell type.

Sex-linked genes exhibit unique patterns of inheritance:

As you just learned, Morgan's discovery of a trait (white eyes) that correlated with the sex of flies was a key episode in the development of the chromosome theory of inheritance. Because the identity of the sex chromosomes in an individual could be inferred by observing the sex of the fly, the behavior of the two members of the pair of sex chromosomes could be correlated with the behavior of the two alleles of the eye color gene. In this section, we'll take a closer look at the role of sex chromosomes in inheritance.

Meiosis in plants

As you learned in Figure 13.6b, meiosis in plants produces spores, not gametes. In flowering plants like the pea, each spore develops into a microscopic haploid gametophyte that contains only a few cells and is located on the parent plant. The gametophyte produces sperm, in pollen grains, and eggs, in the carpel. For simplicity, we will not include the gametophyte stage in our discussion of fertilization in plan

The resulting flies had a much higher proportion of the combinations of traits seen in the P generation flies (called parental phenotypes) than would be expected if the two genes assorted independently. Morgan thus concluded that body color and wing size are usually inherited together in specific (parental) combinations because the genes are linked; they are near each other on the same chromosome: F1 dihybrid female and homozygous recessive male in testcross Most offspring

As you proceed, be sure to keep in mind the distinction between the terms linked genes (two or more genes on the same chromosome that tend to be inherited together) and sex-linked gene (a single gene on a sex chromosome).

Figure 14.15b is a pedigree of the same family, but this time we focus on a recessive trait, the inability of individuals to taste a chemical called PTC (phenylthiocarbamide). Compounds similar to PTC are found in broccoli, brussels sprouts, and related vegetables and account for the bitter taste some people report when eating these foods. We'll use t for the recessive allele and T for the dominant allele, which results in the ability to taste PTC.

As you work your way through the pedigree, notice once again that you can apply what you have learned about Mendelian inheritance to understand the genotypes shown for the family members. An important application of a pedigree is to help us calculate the probability that a future child will have a particular genotype and phenotype.

Independent Assortment of Chromosomes: One aspect of sexual reproduction that generates genetic variation is the random orientation of pairs of homologous chromosomes at metaphase of meiosis I.

At metaphase I, the homologous pairs, each consisting of one maternal and one paternal chromosome, are situated at the metaphase plate. (Note that the terms maternal and paternal refer, respectively, to whether the chromosome in question was contributed by the mother or the father of the individual whose cells are undergoing meiosis.) Each pair may orient with either its maternal or paternal homolog closer to a given pole— its orientation is as random as the flip of a coin. Thus, there is a 50% chance that a particular daughter cell of meiosis I will get the maternal chromosome of a certain homologous pair and a 50% chance that it will get the paternal chromosome.

Genetic Testing and Counseling:

Avoiding simple Mendelian disorders is possible when the risk of a particular genetic disorder can be assessed before a child is conceived or during the early stages of the pregnancy. Many hospitals have genetic counselors who can provide information to prospective parents concerned about a family history for a specific disease. Fetal and newborn testing can also reveal genetic disorders.

As a human develops into a sexually mature adult, mitosis of the zygote and its descendant cells generates all the somatic cells of the body.

Both chromosome sets in the zygote and all the genes they carry are passed with precision to the somatic cells.

Bacteria often respond to environmental change by regulating transcription :

Bacterial cells that can conserve resources and energy have a selective advantage over cells that are unable to do so. Thus, natural selection has favored bacteria that express only the genes whose products are needed by the cell.

RNA Polymerase Binding and Initiation of Transcription: The promoter of a gene includes within it the transcription start point—the nucleotide where RNA polymerase actually begins synthesis of the mRNA—and typically extends several dozen or so nucleotide pairs upstream from the start point (Figure 17.9).

Based on interactions with proteins that will be covered shortly, RNA polymerase binds in a precise location and orientation on the promoter. This in turn determines where transcription starts and which of the two strands of the DNA helix is used as the template.

Using recombination data, Sturtevant and his colleagues were able to map numerous Drosophila genes in linear arrays. They found that the genes clustered into four groups of linked genes (linkage groups). Light microscopy had revealed four pairs of chromosomes in Drosophila, so the linkage map provided additional evidence that genes are located on chromosomes. Each chromosome has a linear array of specific genes, each gene with its own locus (Figure 15.12).

Because a linkage map is based strictly on recombination frequencies, it gives only an approximate picture of a chromosome. The frequency of crossing over is not actually uniform over the length of a chromosome, as Sturtevant assumed, and therefore map units do not correspond to actual physical distances (in nanometers, for instance).

During translation, the sequence of codons along an mRNA molecule is decoded, or translated, into a sequence of amino acids making up a polypeptide chain. The codons are read by the translation machinery in the 5' -> 3' direction along the mRNA. Each codon specifies which one of the 20 amino acids will be incorporated at the corresponding position along a polypeptide.

Because codons are nucleotide triplets, the number of nucleotides making up a genetic message must be three times the number of amino acids in the protein product. For example, it takes 300 nucleotides along an mRNA strand to code for the amino acids in a polypeptide that is 100 amino acids long

Even during interphase, the centromeres and telomeres of chromosomes, as well as other chromosomal regions in some cells, exist in a highly condensed state similar to that seen in a metaphase chromosome. This type of interphase chromatin, visible as irregular clumps with a light microscope, is called heterochromatin, to distinguish it from the less compacted, more dispersed euchromatin ("true chromatin").

Because of its compaction, heterochromatic DNA is largely inaccessible to the machinery in the cell responsible for transcribing the genetic information coded in the DNA, a crucial early step in gene expression. In contrast, the looser packing of euchromatin makes its DNA accessible to this machinery, so the genes present in euchromatin can be transcribed. The chromosome is a dynamic structure that is condensed, loosened, modified, and remodeled as necessary for various cell processes, including mitosis, meiosis, and gene activity. Chemical modifications of histones affect the state of chromatin condensation and also have multiple effects on gene activity, as you'll see in Concept 18.2.

Another example is the mosquito-borne virus called chikungunya (Figure 19.10b), which causes an acute illness with fever, rashes, and persistent joint pain. Chikungunya has long been considered a tropical virus, but it has now appeared in northern Italy and southeastern France. A more recently emerging virus is the Zika virus (Figure 19.10c), which caused an outbreak of disease in spring 2015 in Brazil. Although symptoms of Zika are often mild, the outbreak was noticed because infection of pregnant women was correlated with a striking increase in the number of babies born with abnormally small brains, a condition called microcephaly. Zika is a mosquito-borne flavivirus (like West Nile virus) that infects neural cells, posing a particular danger to fetal brain development.

Because of the neurological defects associated with Zika and its spread to 28 other countries by early 2016, the World Health Organization declared Zika an international health emergency.

In our model, an AABBCC person would be very dark, whereas an aabbcc individual would be very light. An AaBbCc person would have skin of an intermediate shade.

Because the alleles have a cumulative effect, the genotypes AaBbCc and AABbcc would make the same genetic contribution (three units) to skin darkness. There are seven skin color phenotypes that could result from a mating between AaBbCc heterozygotes, as shown in Figure 14.13. In a large number of such matings, the majority of offspring would be expected to have intermediate phenotypes (skin color in the middle range). You can graph the predictions from the Punnett square in the Scientific Skills Exercise. Environmental factors, such as exposure to the sun, also affect the skin color phenotype.

Similar maternal inheritance is also the rule for mitochondrial genes in most animals and plants, because the mitochondria passed on to a zygote come from the cytoplasm of the egg. (The few mitochondria contributed by the sperm appear to be destroyed in the egg by autophagy; see Figure 6.13.)The products of most mitochondrial genes help make up some of the protein complexes of the electron transport chain and ATP synthase (see Figure 9.15). Defects in one or more of these proteins, therefore, reduce the amount of ATP the cell can make and have been shown to cause a number of human disorders in as many as one out of every 5,000 births.

Because the parts of the body most susceptible to energy deprivation are the nervous system and the muscles, most mitochondrial diseases primarily affect these systems. For example, mitochondrial myopathy causes weakness, intolerance of exercise, and muscle deterioration. Another mitochondrial disorder is Leber's hereditary optic neuropathy, which can produce sudden blindness in people as young as their 20s or 30s. The four mutations found thus far to cause this disorder affect oxidative phosphorylation during cellular respiration, a crucial function for the cell (see Concept 9.4).

Genes specify proteins via transcription and translation:

Before going into the details of how genes direct protein synthesis, let's step back and examine how the fundamental relationship between genes and proteins was discovered.

In addition to enabling RNA splicing, eukaryotic host cells are advantageous because many eukaryotic proteins will not function unless they are modified after translation—for example, by the addition of carbohydrate groups (glycosylation) or lipid groups. Bacterial cells cannot carry out these modifications, and if the gene product requiring such processing is from a mammal, even yeast cells may not be able to modify the protein correctly. Several cultured cell types have proved successful as host cells for this purpose, including some mammalian cell lines and an insect cell line that can be infected by a virus (called baculovirus) carrying recombinant DNA.

Besides using vectors, scientists have developed other methods for introducing recombinant DNA into eukaryotic cells. In electroporation, a brief electrical pulse applied to a solution containing cells creates temporary holes in their plasma membranes, through which DNA can enter. (This technique is now commonly used for bacteria as well.) Alternatively, scientists can inject DNA directly into single eukaryotic cells using microscopically thin needles. Another way to get DNA into plant cells is by using the soil bacterium Agrobacterium tumefaciens, as we'll discuss later. Whatever the method, if the introduced DNA is incorporated into a cell's genome by genetic recombination, then it can be expressed by the cell. Expressing different versions of genes in cells allows researchers to study protein function, a topic we'll return to in Concept 20.2.

Analyzing Gene Expression:

Biologists driven to understand the assorted cell types of a multicellular organism, cancer cells, or the developing tissues of an embryo first try to discover which genes are expressed by the cells of interest. The most straightforward way to do this is usually to identify the mRNAs being made. We'll first examine techniques that look for patterns of expression of specific individual genes. Next, we'll explore ways to characterize groups of genes being expressed by cells or tissues of interest. As you will see, all of these procedures depend in some way on base pairing between complementary nucleotide sequences.

Targeting Polypeptides to Specific Locations: In electron micrographs of eukaryotic cells active in protein synthesis, two populations of ribosomes are evident: free and bound (see Figure 6.10). Free ribosomes are suspended in the cytosol and mostly synthesize proteins that stay in the cytosol and function there. In contrast, bound ribosomes are attached to the cytosolic side of the endoplasmic reticulum (ER) or to the nuclear envelope.

Bound ribosomes make proteins of the endomembrane system (see Figure 6.15) as well as proteins secreted from the cell, such as insulin. It is important to note that the ribosomes themselves are identical and can alternate between being free ribosomes one time they are used and being bound ribosomes the next.

Substitution mutations are usually missense mutations; that is, the altered codon still codes for an amino acid and thus makes sense, although not necessarily the right sense.

But a point mutation can also change a codon for an amino acid into a stop codon. This is called a nonsense mutation, and it causes translation to be terminated prematurely; the resulting polypeptide will be shorter than the polypeptide encoded by the normal gene. Most nonsense mutations lead to nonfunctional proteins.

Another variation on dominance relationships between alleles is called codominance; in this variation, the two alleles each affect the phenotype in separate, distinguishable ways. For example, the human MN blood group is determined by codominant alleles for two specific molecules located on the surface of red blood cells, the M and N molecules. A single gene locus, at which two allelic variations are possible, determines the phenotype of this blood group. Individuals homozygous for the M allele (MM) have red blood cells with only M molecules; individuals homozygous for the N allele (NN) have red blood cells with only N molecules.

But both M and N molecules are present on the red blood cells of individuals heterozygous for the M and N alleles (MN). Note that the MN phenotype is not intermediate between the M and N phenotypes, which distinguishes codominance from incomplete dominance. Rather, both M and N phenotypes are exhibited by heterozygotes, since both molecules are present.

Note that if an organism has identical alleles for a particular character then that allele is present in all gametes. Because it is the only allele that can be passed on to offspring, the offspring always look like their parents; this explains why these plants are true-breeding.

But if different alleles are present, as in the F1 hybrids, then 50% of the gametes receive the dominant allele and 50% receive the recessive allele.

Scientists remained skeptical, however, since many still viewed proteins as better candidates for the genetic material. Also, many biologists were not convinced that bacterial genes would be similar in composition and function to those of more complex organisms.

But the major reason for the continued doubt was that so little was known about DNA.

In time, you'll be able to solve genetics problems faster by using the rules of probability than by filling in Punnett squares. We cannot predict with certainty the exact numbers of progeny of different genotypes resulting from a genetic cross.

But the rules of probability give us the likelihood of various outcomes. Usually, the larger the sample size, the closer the results will conform to our predictions. Mendel understood this statistical feature of inheritance and had a keen sense of the rules of chance. It was for this reason that he set up his experiments so as to generate, and then count, large numbers of offspring from his crosses.

:Epistasis: In epistasis (from the Greek for "standing upon"), the phenotypic expression of a gene at one locus alters that of a gene at a second locus. An example will help clarify this concept. In Labrador retrievers (commonly called "Labs"), black coat color is dominant to brown. Let's designate B and b as the two alleles for this character. For a Lab to have brown fur, its genotype must be bb; these dogs are called chocolate Labs.

But there is more to the story. A second gene determines whether or not pigment will be deposited in the hair. The dominant allele, symbolized by E, results in the deposition of either black or brown pigment, depending on the genotype at the first locus. But if the Lab is homozygous recessive for the second locus (ee), then the coat is yellow, regardless of the genotype at the black/brown locus (yellow Labs). In this case, the gene for pigment deposition (E/e) is said to be epistatic to the gene that codes for black or brown pigment (B/b).

The nitrogenous bases of the double helix are paired in specific combinations: adenine (A) with thymine (T), and guanine (G) with cytosine (C). It was mainly by trial and error that Watson and Crick arrived at this key feature of DNA. At first, Watson imagined that the bases paired like with like—for example, A with A and C with C.

But this model did not fit the X-ray data, which suggested that the double helix had a uniform diameter.

segregation and independent assortment. As you can see in Figure 15.2, the separation of homologs during anaphase I accounts for the segregation of the two alleles of a gene into separate gametes, and the random arrangement of chromosome pairs at metaphase I accounts for independent assortment of the alleles for two or more genes located on different homolog pairs. This figure traces the same dihybrid pea cross you learned about in Figure 14.8.

By carefully studying Figure 15.2, you can see how the behavior of chromosomes during meiosis in the F1 generation and subsequent random fertilization give rise to the F2 phenotypic ratio observed by Mendel.

In the 1940s, scientists began using the genetic approach— the study of mutants—to investigate Drosophila development. That approach has had spectacular success. These studies have established that genes control development and have led to an understanding of the key roles that specific molecules play in defining position and directing differentiation.

By combining anatomical, genetic, and biochemical approaches to the study of Drosophila development, researchers have discovered developmental principles common to many other species, including humans.

Building a Structural Model of DNA: Scientific Inquiry Once most biologists were convinced that DNA was the genetic material, the challenge was to determine how the structure of DNA could account for its role in inheritance.

By the early 1950s, the arrangement of covalent bonds in a nucleic acid polymer was well established (see Figure 16.5), and researchers focused on discovering the three-dimensional structure of DNA. Among the scientists working on the problem were Linus Pauling, at the California Institute of Technology, and Maurice Wilkins and Rosalind Franklin, at King's College in London. First to come up with the complete answer, however, were two scientists who were relatively unknown at the time—the American James Watson and the Englishman Francis Crick.

Medical scientists have also developed methods for isolating fetal cells, or even fetal DNA, that have escaped into the mother's blood. Although very few are present, the cells can be cultured and tested, and the fetal DNA can be analyzed. In 2012, researchers were able to analyze the entire genome of a fetus, comparing sequences of samples obtained from both parents and fetal DNA found in the mother's blood.

Cell-free fetal DNA tests and other blood tests are increasingly being used as noninvasive prenatal screening tests for certain disorders; a positive result indicates to the parents that further diagnostic testing, such as amniocentesis or CVS, should be considered. Imaging techniques allow a physician to examine a fetus directly for major anatomical abnormalities that might not show up in genetic tests. In the ultrasound technique, for example, reflected sound waves are used to produce an image of the fetus by a simple noninvasive procedure.

Unlike a gamete, a haploid spore doesn't fuse with another cell but divides mitotically, generating a multicellular haploid stage called the gametophyte.

Cells of the gametophyte give rise to gametes by mitosis. Fusion of two haploid gametes at fertilization results in a diploid zygote, which develops into the next sporophyte generation. Therefore, in this type of life cycle, the sporophyte generation produces a gametophyte as its offspring, and the gametophyte generation produces the next sporophyte generation (see Figure 13.6b). The term alternation of generations fits well as a name for this type of life cycle.

Although the ribosomes of bacteria and eukaryotes are very similar in structure and function, eukaryotic ribosomes are slightly larger, as well as differing somewhat from bacterial ribosomes in their molecular composition. The differences are medically significant.

Certain antibiotic drugs can inactivate bacterial ribosomes without affecting eukaryotic ribosomes. These drugs, including tetracycline and streptomycin, are used to combat bacterial infections.

The combination of crossing over and sister chromatid cohesion along the arms results in the formation of a chiasma.

Chiasmata hold homologs together as the spindle forms for the first meiotic division. At the onset of anaphase I, the release of cohesion along sister chromatid arms allows homologs to separate.

Newborn Screening: Some genetic disorders can be detected at birth by simple biochemical tests that are now routinely performed in most hospitals in the United States. One common screening program is for phenylketonuria (PKU), a recessively inherited disorder that occurs in about one out of every 10,000-15,000 births in the United States.

Children with this disease cannot properly metabolize the amino acid phenylalanine. This compound and its by-product, phenylpyruvate, can accumulate to toxic levels in the blood, causing severe intellectual disability (mental retardation). However, if PKU is detected in the newborn, a special diet low in phenylalanine will usually allow normal development. (Among many other substances, this diet excludes the artificial sweetener aspartame, which contains phenylalanine.) Unfortunately, few other genetic disorders are treatable at present.

The Genetic Code: When biologists began to suspect that the instructions for protein synthesis were encoded in DNA, they recognized a problem: There are only four nucleotide bases to specify 20 amino acids. Thus, the genetic code cannot be a language like Chinese, where each written symbol corresponds to a word. How many nucleotides, then, would turn out to correspond to an amino acid?

Codons: Triplets of Nucleotides: If each kind of nucleotide base were translated into an amino acid, only four amino acids could be specified, one per nucleotide base. Would a language of two-letter code words suffice? The two-nucleotide sequence AG, for example, could specify one amino acid, and GT could specify another. Since there are four possible nucleotide bases in each position, this would give us 16 (that is, 4 * 4, or 42 ) possible arrangements—still not enough to code for all 20 amino acids.

Despite being on the same chromosome and thus being physically connected, the genes are genetically unlinked; alleles of such genes assort independently, as if they were on different chromosomes. In fact, at least two of the genes for pea characters that Mendel studied are now known to be on the same chromosome, but the distance between them is so great that linkage is not observed in genetic crosses.

Consequently, the two genes behaved as if they were on different chromosomes in Mendel's experiments. Genes located far apart on a chromosome are mapped by adding the recombination frequencies from crosses involving closer pairs of genes lying between the two distant genes.

One example involves a gene called Pax-6, which has been found in animals as diverse as vertebrates and fruit flies. The vertebrate Pax-6 gene product (the PAX-6 protein) triggers a complex program of gene expression resulting in formation of the vertebrate eye, which has a single lens. Expression of the fly Pax-6 gene leads to formation of the compound fly eye, which is quite different from the vertebrate eye. When the mouse Pax-6 gene was cloned and introduced into a fly embryo so that it replaced the fly's own Pax-6 gene, researchers were surprised to see that the mouse version of the gene led to formation of a compound fly eye (see Figure 50.16).

Conversely, when the fly Pax-6 gene was transferred into a vertebrate embryo—a frog, in this case—a frog eye formed. Although the genetic programs triggered in vertebrates and flies generate very different eyes, the two versions of the Pax-6 gene can substitute for each other to trigger lens development, evidence of their evolution from a gene in a very ancient common ancestor. Because of their ancient evolutionary roots, all living organisms share the same basic mechanisms of gene expression. This commonality is the basis of many recombinant DNA techniques described in this chapter.

Many signaling molecules, such as nonsteroid hormones and growth factors, bind to receptors on a cell's surface and never actually enter the cell. Such molecules can control gene expression indirectly by triggering signal transduction pathways that activate particular transcription factors (see Figure 11.15).

Coordinate regulation in such pathways is the same as for steroid hormones: Genes with the same sets of control elements are activated by the same chemical signals. Because this system for coordinating gene regulation is so widespread, biologists think that it probably arose early in evolutionary history.

A likely scenario for the 1918 pandemic and others is that the virus mutated as it passed from one host species to another. When an animal like a pig or a bird is infected with more than one strain of flu virus, the different strains can undergo genetic recombination if the RNA molecules making up their genomes mix and match during viral assembly. Pigs were probably the main hosts for recombination that led to the 2009 flu virus, which turns out to contain sequences from bird, pig, and human flu viruses.

Coupled with mutation, these reassortments can lead to the emergence of a viral strain capable of infecting human cells. People who have never been exposed to that particular strain before will lack immunity, and the recombinant virus has the potential to be highly pathogenic. If such a flu virus recombines with viruses that circulate widely among humans, it may acquire the ability to spread easily from person to person, dramatically increasing the potential for a major human outbreak.

Using improved techniques of microscopy, cytologists worked out the process of mitosis in 1875 and meiosis in the 1890s. (See the drawing of mitosis published by German biologist Walther Flemming in 1882.)

Cytology and genetics converged as biologists began to see parallels between the behavior of Mendel's proposed hereditary factors during sexual life cycles and the behavior of chromosomes: As shown in Figure 15.2, chromosomes and genes are both present in pairs in diploid cells, and homologous chromosomes separate and alleles segregate during the process of meiosis. After meiosis, fertilization restores the paired condition for both chromosomes and genes. Around 1902, Walter S. Sutton, Theodor Boveri, and others independently noted these parallels and began to develop the chromosome theory of inheritance. According to this theory, Mendelian genes have specific loci (positions) along chromosomes, and it is the chromosomes that undergo segregation and independent assortment.

Given the extensive functions of ncRNAs, it is not surprising that many of the ncRNAs characterized thus far play important roles in embryonic development—the topic we turn to in the next section.

Embryonic development is perhaps the ultimate example of precisely regulated gene expression.

The Evolutionary Significance of Genetic Variation Within Populations: Now that you've learned how new combinations of genes arise among offspring in a sexually reproducing population, how does the genetic variation in a population relate to evolution?

Darwin recognized that a population evolves through the differential reproductive success of its variant members. On average, those individuals best suited to the local environment leave the most offspring, thereby transmitting their genes. Thus, natural selection results in the accumulation of genetic variations favored by the environment.

Evolution of the Genetic Code: The genetic code is nearly universal, shared by organisms from the simplest bacteria to the most complex plants and animals. The mRNA codon CCG, for instance, is translated as the amino acid proline in all organisms whose genetic code has been examined. In laboratory experiments, genes can be transcribed and translated after being transplanted from one species to another, sometimes with quite striking results, as shown in Figure 17.7. Bacteria can be programmed by the insertion of human genes to synthesize certain human proteins for medical use, such as insulin. Such applications have produced many exciting developments in the area of biotechnology (see Concept 20.4).

Despite a small number of exceptions, the evolutionary significance of the code's near universality is clear. A language shared by all living things must have been operating very early in the history of life—early enough to be present in the common ancestor of all present-day organisms. A shared genetic vocabulary is a reminder of the kinship of all life.

Despite its speed and specificity, PCR amplification cannot substitute for gene cloning in cells to make large amounts of a gene. This is because occasional errors during PCR replication limit the number of good copies and the length of DNA fragments that can be copied. Instead, PCR is used to provide the specific DNA fragment for cloning. PCR primers are synthesized to include a restriction site at each end of the DNA fragment that matches the site in the cloning vector, and the fragment and vector are cut and ligated together (Figure 20.8). The resulting plasmids are sequenced so that those with error-free inserts can be selected.

Devised in 1985, PCR has had a major impact on biological research and genetic engineering. PCR has been used to amplify DNA from a wide variety of sources: a 40,000-year-old frozen woolly mammoth (see the photo on the first page of this chapter); fingerprints or tiny amounts of blood, tissue, or semen found at crime scenes; single embryonic cells for rapid prenatal diagnosis of genetic disorders (see Figure 14.19); and cells infected with viruses that are difficult to detect, such as HIV. (To test for HIV, viral genes are amplified.) We'll return to applications of PCR later.

The structure of a ribosome reflects its function of bringing mRNA together with tRNAs carrying amino acids. In addition to a binding site for mRNA, each ribosome has three binding sites for tRNA (Figure 17.18). The P site (peptidyltRNA binding site) holds the tRNA carrying the growing polypeptide chain, while the A site (aminoacyl-tRNA binding site) holds the tRNA carrying the next amino acid to be added to the chain.

Discharged tRNAs leave the ribosome from the E site (exit site). The ribosome holds the tRNA and mRNA in close proximity and positions the new amino acid so that it can be added to the carboxyl end of the growing polypeptide. It then catalyzes the formation of the peptide bond. As the polypeptide becomes longer, it passes through an exit tunnel in the ribosome's large subunit. When the polypeptide is complete, it is released through the exit tunnel.

Viruses and prions are formidable pathogens in animals and plants:

Diseases caused by viral infections afflict humans, agricultural crops, and livestock worldwide. Other smaller, less complex entities known as prions also cause disease in animals. We'll first discuss animal viruses.

Extending Mendelian Genetics for Two or More Genes:

Dominance relationships, multiple alleles, and pleiotropy all have to do with the effects of the alleles of a single gene. We now consider two situations in which two or more genes are involved in determining a particular phenotype. In the first case, called epistasis, one gene affects the phenotype of another because the two gene products interact; in the second case, called polygenic inheritance, multiple genes independently affect a single trait.

Most of the genes carried on the X chromosome do not have counterparts on the tiny Y, and the Y chromosome has genes lacking on the X.

Due to their role in sex determination, the X and Y chromosomes are called sex chromosomes. The other chromosomes are called autosomes.

A Genetic Program for Embryonic Development: The photos in Figure 18.16 illustrate the dramatic difference between a frog zygote (fertilized egg) and the tadpole it becomes. This remarkable transformation results from three interrelated processes: cell division, cell differentiation, and morphogenesis. Through a succession of mitotic cell divisions, the zygote gives rise to a large number of cells. Cell division alone, however, would merely produce a great ball of identical cells, nothing like a tadpole.

During embryonic development, cells not only increase in number, but also undergo cell differentiation, the process by which cells become specialized in structure and function. Moreover, the different kinds of cells are not randomly distributed but are organized into tissues and organs in a particular threedimensional arrangement. The physical processes that give an organism its shape constitute morphogenesis, the development of the form of an organism and its structures. All three processes are rooted in cellular behavior.

Does Mendel's segregation model account for the 3:1 ratio he observed in the F2 generation of his numerous crosses? For the flower-color character, the model predicts that the two different alleles present in an F1 individual will segregate into gametes such that half the gametes will have the purple-flower allele and half will have the white-flower allele.

During selfpollination, gametes of each class unite randomly. An egg with a purple-flower allele has an equal chance of being fertilized by a sperm with a purple-flower allele or by one with a white-flower allele. Since the same is true for an egg with a white-flower allele, there are four equally likely combinations of sperm and egg.

These cues tell a cell its location relative to the body axes and to neighboring cells and determine how the cell and its descendants will respond to future molecular signals.

During the first half of the 20th century, classical embryologists made detailed anatomical observations of embryonic development in a number of species and performed experiments in which they manipulated embryonic tissues. Although this research laid the groundwork for understanding the mechanisms of development, it did not reveal the specific molecules that guide development or determine how patterns are established.

Nucleotide sequences that affect how long an mRNA remains intact are often found in the untranslated region at the 3′ end of the molecule (see Figure 18.8). In one experiment, researchers transferred such a sequence from the short-lived mRNA for a growth factor to the 3′ end of a normally stable globin mRNA. The globin mRNA was quickly degraded.

During the past few years, other mechanisms that degrade or block expression of mRNA molecules have come to light. They involve a group of newly discovered RNA molecules that regulate gene expression at several levels, as we'll discuss shortly

Breast Cancer Subtypes

Each breast cancer subtype is characterized by the overexpression (indicated by ++ or +++) or absence (-) of three signal receptors: ERα, PR, and HER2. Breast cancer treatments are becoming more effective because they can be tailored to the specific cancer subtype. Luminal A: • ERα+++ • PR++ • HER2- • 40% of breast cancers • Best prognosis Luminal B: • ERα++ • PR++ • HER2- (shown here); some HER2++ • 15-20% of breast cancers • Poorer prognosis than luminal A subtype Basal-like HER2 Treatment with Herceptin for the HER2 subtype Both luminal subtypes overexpress ERα (luminal A more than luminal B) and PR, and usually lack expression of HER2. Both can be treated with drugs that target ERα and inactivate it, the most well-known drug being tamoxifen. These subtypes can also be treated with drugs that inhibit estrogen synthesis.

Stretched out, the DNA of an E. coli cell would measure about a millimeter in length, which is 500 times longer than the cell. Within a bacterium, however, certain proteins cause the chromosome to coil and "supercoil," densely packing it so that it fills only part of the cell. Unlike the nucleus of a eukaryotic cell, this dense region of DNA in a bacterium, called the nucleoid, is not bounded by membrane (see Figure 6.5).

Each eukaryotic chromosome contains a single linear DNA double helix that, in humans, averages about 1.5 * 108 nucleotide pairs. This is an enormous amount of DNA relative to a chromosome's condensed length. If completely stretched out, such a DNA molecule would be about 4 cm long, thousands of times the diameter of a cell nucleus—and that's not even considering the DNA of the other 45 human chromosomes!

Most DNA polymerases require a primer and a DNA template strand, along which complementary DNA nucleotides are lined up. In E. coli, DNA polymerase III (abbreviated DNA pol III) adds a DNA nucleotide to the RNA primer and then continues adding DNA nucleotides, complementary to the parental DNA template strand, to the growing end of the new DNA strand. The rate of elongation is about 500 nucleotides per second in bacteria and 50 per second in human cells.

Each nucleotide to be added to a growing DNA strand consists of a sugar attached to a base and to three phosphate groups. You have already encountered such a molecule—ATP (adenosine triphosphate; see Figure 8.9). The only difference between the ATP of energy metabolism and dATP, the adenine nucleotide used to make DNA, is the sugar component, which is deoxyribose in the building block of DNA but ribose in ATP. Like ATP, the nucleotides used for DNA synthesis are chemically reactive, partly because their triphosphate tails have an unstable cluster of negative charge. DNA polymerase catalyzes the addition of each monomer via a dehydration reaction (see Figure 5.2a). As each monomer is joined to the growing end of a DNA strand, two phosphate groups are lost as a molecule of pyrophosphate ( P — P i ). Subsequent hydrolysis of the pyrophosphate to two molecules of inorganic phosphate ( P i ) is a coupled exergonic reaction that helps drive the polymerization reaction (Figure 16.14).

Other advantages of using peas are their short generation time and the large number of offspring from each mating. Furthermore, Mendel could strictly control mating between plants (Figure 14.2).

Each pea flower has both pollen-producing organs (stamens) and an egg-bearing organ (carpel). In nature, pea plants usually self-fertilize: Pollen grains from the stamens land on the carpel of the same flower, and sperm released from the pollen grains fertilize eggs present in the carpel.

It is customary to describe the flow of information from gene to protein in linguistic terms. Just as specific sequences of letters communicate information in a language such as English, both nucleic acids and proteins are polymers with specific sequences of monomers that convey information. In DNA or RNA, the monomers are the four types of nucleotides, which differ in their nitrogenous bases. Genes are typically hundreds or thousands of nucleotides long, each gene having a specific sequence of nucleotides.

Each polypeptide of a protein also has monomers arranged in a particular linear order (the protein's primary structure; see Figure 5.18), but its monomers are amino acids. Thus, nucleic acids and proteins contain information written in two different chemical languages. Getting from DNA to protein requires two major stages: transcription and translation.

To achieve cross-pollination of two plants, Mendel removed the immature stamens of a plant before they produced pollen and then dusted pollen from another plant onto the altered flowers (see Figure 14.2).

Each resulting zygote then developed into a plant embryo encased in a seed (pea). His method allowed Mendel to always be sure of the parentage of new seeds.

Coordinately Controlled Genes in Eukaryotes: How does the eukaryotic cell deal with a group of genes of related function that need to be turned on or off at the same time?

Earlier in this chapter, you learned that in bacteria, such coordinately controlled genes are often clustered into an operon, which is regulated by a single promoter and transcribed into a single mRNA molecule. Thus, the genes are expressed together, and the encoded proteins are produced concurrently. With a few exceptions, operons that work in this way have not been found in eukaryotic cells.

Mutations of one or a few nucleotides can affect protein structure and function: Now that you have explored the process of gene expression, you are ready to understand the effects of changes to the genetic information of a cell. These changes, called mutations, are responsible for the huge diversity of genes found among organisms because mutations are the ultimate source of new genes.

Earlier, we considered chromosomal rearrangements that affect long segments of DNA (see Figure 15.14); these are considered large-scale mutations. Here we examine small-scale mutations of one or a few nucleotide pairs, including point mutations, changes in a single nucleotide pair of a gene

Evolution of Viruses: We began this chapter by asking whether or not viruses are alive. Viruses do not really fit our definition of living organisms. An isolated virus is biologically inert, unable to replicate its genes or regenerate its own ATP. Yet it has a genetic program written in the universal language of life. Do we think of viruses as nature's most complex associations of molecules or as the simplest forms of life?

Either way, we must bend our usual definitions. Although viruses cannot replicate or carry out metabolic activities independently, their use of the genetic code makes it hard to deny their evolutionary connection to the living world.

Elongation of the Polypeptide Chain: In the elongation stage of translation, amino acids are added one by one to the previous amino acid at the C-terminus of the growing chain. Each addition involves several proteins called elongation factors and occurs in a three-step cycle described in Figure 17.20.

Energy expenditure occurs in the first and third steps. Codon recognition requires hydrolysis of one molecule of GTP, which increases the accuracy and efficiency of this step. One more GTP is hydrolyzed to provide energy for the translocation step.

Eukaryotic cells modify RNA after transcription:

Enzymes in the eukaryotic nucleus modify pre-mRNA in specific ways before the genetic message is dispatched to the cytoplasm. During this RNA processing, both ends of the primary transcript are altered. Also, in most cases, certain interior sections of the RNA molecule are cut out and the remaining parts spliced together. These modifications produce an mRNA molecule ready for translation.

Alterations of Chromosome Structure:

Errors in meiosis or damaging agents such as radiation can cause breakage of a chromosome, which can lead to four types of changes in chromosome structure (Figure 15.14) deletion, duplication, inversion, translocation.

Most prokaryotes have a circular chromosome, with no ends, so the shortening of DNA does not occur. But what protects the genes of linear eukaryotic chromosomes from being eroded away during successive rounds of DNA replication?

Eukaryotic chromosomal DNA molecules have special nucleotide sequences called telomeres at their ends (Figure 16.21). Telomeres do not contain genes; instead, the DNA typically consists of multiple repetitions of one short nucleotide sequence. In each human telomere, for example, the six-nucleotide sequence TTAGGG is repeated between 100 and 1,000 times.

Replicating the Ends of DNA Molecules: For linear DNA, such as the DNA of eukaryotic chromosomes, the usual replication machinery cannot complete the 5′ ends of daughter DNA strands. (This is another consequence of the fact that a DNA polymerase can add nucleotides only to the 3′ end of a preexisting polynucleotide.)

Even if an Okazaki fragment can be started with an RNA primer hydrogen-bonded to the very end of the template strand, once that primer is removed, it cannot be replaced with DNA because there is no 3′ end available for nucleotide addition (Figure 16.20). As a result, repeated rounds of replication produce shorter and shorter DNA molecules with uneven ("staggered") ends.

Recessive homozygotes could also result from Aa * aa and aa * aa matings, but if the disorder is lethal before reproductive age or results in sterility (neither of which is true for albinism), no aa individuals will reproduce.

Even if recessive homozygotes are able to reproduce, such matings will occur relatively rarely because these individuals account for a much smaller percentage of the population than heterozygous carriers (for reasons we'll examine in Concept 23.2).

Split Genes and RNA Splicing: A remarkable stage of RNA processing in the eukaryotic nucleus is RNA splicing (Figure 17.12), where large portions of the RNA molecules are removed and the remaining portions are reconnected. This cut-and-paste job is similar to editing a movie. The average length of a transcription unit along a human DNA molecule is about 27,000 nucleotide pairs, so the primary RNA transcript is also that long. However, the average-sized protein of 400 amino acids requires only 1,200 nucleotides in RNA to code for it. (Remember, each amino acid is encoded by a triplet of nucleotides.) This is because most eukaryotic genes and their RNA transcripts have long noncoding stretches of nucleotides, regions that are not translated.

Even more surprising is that most of these noncoding sequences are interspersed between coding segments of the gene and thus between coding segments of the pre-mRNA. In other words, the sequence of DNA nucleotides that codes for a eukaryotic polypeptide is usually not continuous; it is split into segments. The noncoding segments of nucleic acid that lie between coding regions are called intervening sequences, or introns. The other regions are called exons, because they are eventually expressed, usually by being translated into amino acid sequences. (Exceptions include the UTRs of the exons at the ends of the RNA, which make up part of the mRNA but are not translated into protein. Because of these exceptions, you may prefer to think of exons as sequences of RNA that exit the nucleus.) The terms intron and exon are used for both RNA sequences and the DNA sequences that specify them.

In this section, we have seen how a carefully orchestrated program of sequential gene regulation controls the transformation of a fertilized egg into a multicellular organism. The program is carefully balanced between turning on the genes for differentiation in the right place and turning off other genes.

Even when an organism is fully developed, gene expression is regulated in a similarly fine-tuned manner. In the final section of the chapter, we'll consider how fine this tuning is by looking at how specific changes in expression of just a few genes can lead to the development of cancer.

Muscle cells develop from embryonic precursor cells that have the potential to develop into a number of cell types, including cartilage cells and fat cells, but particular conditions commit them to becoming muscle cells. Although the committed cells appear unchanged under the microscope, determination has occurred, and they are now myoblasts.

Eventually, myoblasts start to churn out large amounts of muscle-specific proteins and fuse to form mature, elongated, multinucleate skeletal muscle cells.

Coordinate control of dispersed genes in a eukaryotic cell often occurs in response to chemical signals from outside the cell. A steroid hormone, for example, enters a cell and binds to a specific intracellular receptor protein, forming a hormone-receptor complex that serves as a transcription activator (see Figure 11.9).

Every gene whose transcription is stimulated by a given steroid hormone, regardless of its chromosomal location, has a control element recognized by that hormone-receptor complex. This is how estrogen activates a group of genes that stimulate cell division in uterine cells, preparing the uterus for pregnancy.

Consider, for example, the mouse gene for insulin-like growth factor 2 (Igf2), one of the first imprinted genes to be identified. Although this growth factor is required for normal prenatal growth, only the paternal allele is expressed (Figure 15.17a).

Evidence that the Igf2 gene is imprinted came initially from crosses between normal-sized (wild-type) mice and dwarf (mutant) mice homozygous for a recessive mutation in the Igf2 gene. The phenotypes of heterozygous offspring (with one normal allele and one mutant) differed depending on whether the mutant allele came from the mother or the father (Figure 15.17b).

In the S phase of the cell cycle, the centromeric regions of DNA must be loosened for chromosomal replication and then re-condensed into heterochromatin in preparation for mitosis. In some yeasts, siRNAs produced by the yeast cells themselves are required to re-form the heterochromatin at the centromeres. A model for how this happens is shown in Figure 18.15.

Exactly how the process starts is still under debate, but biologists agree on the general idea: The siRNA system in yeast interacts with other ncRNAs and with chromatin-modifying enzymes to condense the centromere chromatin into heterochromatin. Under most conditions in mammalian cells, siRNAs are not known to occur, and the mechanism for centromere DNA condensation is not yet understood. However, it may turn out to involve other small ncRNAs.

Evolutionary Significance of Altered DNA Nucleotides:

Faithful replication of the genome and repair of DNA damage are important for the functioning of the organism and for passing on a complete, accurate genome to the next generation. The error rate after proofreading and repair is extremely low, but rare mistakes do slip through. Once a mismatched nucleotide pair is replicated, the sequence change is permanent in the daughter molecule that has the incorrect nucleotide as well as in any subsequent copies. As we mentioned earlier, a permanent change in the DNA sequence is called a mutation.

In general, the steps of the human life cycle are typical of many sexually reproducing animals. Indeed, the processes of fertilization and meiosis are also the hallmarks of sexual reproduction in plants, fungi, and protists just as in animals.

Fertilization and meiosis alternate in sexual life cycles, maintaining a constant number of chromosomes in each species from one generation to the next

Figure 15.10 shows how crossing over in a dihybrid female fly resulted in recombinant eggs and ultimately recombinant offspring in Morgan's testcross. Most eggs had a chromosome with either the b+ vg+ or b vg parental genotype, but some had a recombinant chromosome (b+ vg or b vg+ )

Fertilization of all classes of eggs by homozygous recessive sperm (b vg) produced an offspring population in which 17% exhibited a nonparental, recombinant phenotype, reflecting combinations of alleles not seen before in either P generation parent. In the Scientific Skills Exercise, you can use a statistical test to analyze the results from an F1 dihybrid testcross and see whether the two genes assort independently or are linked.

As a result of meiosis, each human sperm and egg is haploid (n = 23).

Fertilization restores the diploid condition by combining two sets of chromosomes, and the human life cycle is repeated, generation after generation (see Figure 13.5).

Figure 18.6 summarizes the process of gene expression in a eukaryotic cell, highlighting key stages in the expression of a protein-coding gene. Each stage depicted in Figure 18.6 is a potential control point at which gene expression can be turned on or off, accelerated, or slowed down.

Fifty or so years ago, an understanding of the mechanisms that control gene expression in eukaryotes seemed almost hopelessly out of reach. Since then, new research methods, notably advances in DNA technology (see Chapter 20), have enabled molecular biologists to uncover many details of eukaryotic gene regulation. In all organisms, gene expression is commonly controlled at transcription; regulation at this stage often occurs in response to signals coming from outside the cell, such as hormones or other signaling molecules. For this reason, the term gene expression is often equated with transcription for both bacteria and eukaryotes. While this may often be the case for bacteria, the greater complexity of eukaryotic cell structure and function provides opportunities for regulating gene expression at many additional stages (see Figure 18.6). In the remainder of this section, we'll examine some of the important control points of eukaryotic gene expression more closely

Pedigree Analysis: Unable to manipulate the mating patterns of people, geneticists instead analyze the results of matings that have already occurred. They do so by collecting information about a family's history for a particular trait and assembling this information into a family tree describing the traits of parents and children across the generations—a family pedigree.

Figure 14.15a shows a three-generation pedigree that traces the occurrence of a pointed contour of the hairline on the forehead. This trait, called a widow's peak, is due to a dominant allele, W. Because the widow's-peak allele is dominant, all individuals who lack a widow's peak must be homozygous recessive (ww). The two grandparents with widow's peaks must have the Ww genotype, since some of their offspring are homozygous recessive. The offspring in the second generation who do have widow's peaks must also be heterozygous, because they are the products of Ww * ww matings. The third generation in this pedigree consists of two sisters. The one who has a widow's peak could be either homozygous (WW) or heterozygous (Ww), given what we know about the genotypes of her parents (both Ww).

British geneticist Mary Lyon demonstrated that the selection of which X chromosome will form the Barr body occurs randomly and independently in each embryonic cell present at the time of X inactivation. As a consequence, females consist of a mosaic of two types of cells: those with the active X derived from the father and those with the active X derived from the mother. After an X chromosome is inactivated in a particular cell, all mitotic descendants of that cell have the same inactive X. Thus, if a female is heterozygous for a sex-linked trait, about half of her cells will express one allele, while the others will express the alternate allele.

Figure 15.8 shows how this mosaicism results in the patchy coloration of a tortoiseshell cat. In humans, mosaicism can be observed in a recessive X-linked mutation that prevents the development of sweat glands. A woman who is heterozygous for this trait has patches of normal skin and patches of skin lacking sweat glands. Inactivation of an X chromosome involves modification of the DNA and proteins bound to it called histones, including attachment of methyl groups (—CH3) to DNA nucleotides.

Synthesis of the leading strand and synthesis of the lagging strand occur concurrently and at the same rate. The lagging strand is so named because its synthesis is delayed slightly relative to synthesis of the leading strand; each new fragment of the lagging strand cannot be started until enough template has been exposed at the replication fork

Figure 16.17 and Table 16.1 summarize DNA replication. Please study them carefully before proceeding.

What is the molecular basis for muscle cell differentiation? The MyoD protein is a transcription factor (see Figure 18.9) that binds to specific control elements in the enhancers of various target genes and stimulates their expression (Figure 18.18). Some target genes for MyoD encode still other muscle-specific transcription factors. MyoD also stimulates expression of the myoD gene itself, an example of positive feedback that perpetuates MyoD's effect in maintaining the cell's differentiated state. Presumably, all the genes activated by MyoD have enhancer control elements recognized by MyoD and are thus coordinately controlled.

Finally, the secondary transcription factors activate the genes for proteins such as myosin and actin that confer the unique properties of skeletal muscle cells. We have now seen how different programs of gene expression that are activated in the fertilized egg can result in differentiated cells and tissues. But for the tissues to function effectively in the organism as a whole, the organism's body plan—its overall three-dimensional arrangement—must be established and superimposed on the differentiation process. Next we'll investigate the molecular basis for the establishment of the body plan, using the well-studied fruit fly Drosophila melanogaster as an example.

Note that in a cross involving heterozygous and homozygous allele pairs (for example, Yy * yy), the probability of heterozygous offspring is 1 ⁄ 2 and the probability of homozygous offspring is 1 ⁄ 2.

Finally, we use the addition rule to add the probabilities for all the different genotypes that fulfill the condition of at least two recessive traits resulting from our PpYyRr * Ppyyrr cross, as shown below: View page 278

Mendel's Model: Mendel developed a model to explain the 3:1 inheritance pattern that he consistently observed among the F2 offspring in his pea experiments. We describe four related concepts making up this model, the fourth of which is the law of segregation.

First, alternative versions of genes account for variations in inherited characters. The gene for flower color in pea plants, for example, exists in two versions, one for purple flowers and the other for white flowers. These alternative versions of a gene are called alleles.Today, we can relate this concept to chromosomes and DNA. As shown in Figure 14.4, each gene is a sequence of nucleotides at a specific place, or locus, along a particular chromosome. he DNA at that locus, however, can vary slightly in its nucleotide sequence. This variation in information content can affect the function of the encoded protein and thus an inherited character of the organism.

Bacterial Defenses Against Phages: After reading about the lytic cycle, you may have wondered why phages haven't exterminated all bacteria. Lysogeny is one major reason why bacteria have been spared from extinction caused by phages. Bacteria also have their own defenses against phages.

First, natural selection favors bacterial mutants with surface proteins that are no longer recognized as receptors by a particular type of phage. Second, when phage DNA does enter a bacterium, the DNA often is identified as foreign and cut up by cellular enzymes called restriction enzymes, which are so named because they restrict a phage's ability to replicate within the bacterium. (Restriction enzymes are used in molecular biology and DNA cloning techniques; see Concept 20.1.) The bacterium's own DNA is methylated in a way that prevents attack by its own restriction enzymes. A third defense is a system present in both bacteria and archaea called the CRISPR-Cas system.

A repressor protein is encoded by a regulatory gene—in this case, a gene called trpR; trpR is located some distance from the trp operon and has its own promoter. Regulatory genes are expressed continuously, although at a low rate, and a few trp repressor molecules are always present in E. coli cells. Why, then, is the trp operon not switched off permanently?

First, the binding of repressors to operators is reversible. An operator alternates between two states: one with the repressor bound and one without the repressor bound. The relative duration of the repressor-bound state is higher when more active repressor molecules are present. Second, the trp repressor, like most regulatory proteins, is an allosteric protein, with two alternative shapes: active and inactive (see Figure 8.20). The trp repressor is synthesized in the inactive form, which has little affinity for the trp operator. Only when a tryptophan molecule binds to the trp repressor at an allosteric site does the repressor protein change to the active form that can attach to the operator, turning the operon off

Alteration of mRNA Ends: Each end of a pre-mRNA molecule is modified in a particular way (Figure 17.11). The 5' end, which is synthesized first, receives a 5' cap, a modified form of a guanine (G) nucleotide added onto the 5' end after transcription of the first 20-40 nucleotides. The 3' end of the pre-mRNA molecule is also modified before the mRNA exits the nucleus. Recall that the pre-mRNA is cut and released soon after the polyadenylation signal, AAUAAA, is transcribed. At the 3' end, an enzyme then adds 50-250 more adenine (A) nucleotides, forming a poly-A tail. The 5' cap and poly-A tail share several important functions.

First, they seem to facilitate the export of the mature mRNA from the nucleus. Second, they help protect the mRNA from degradation by hydrolytic enzymes. And third, they help ribosomes attach to the 5' end of the mRNA once the mRNA reaches the cytoplasm. Figure 17.11 also shows the untranslated regions (UTRs) at the 5' and 3' ends of the mRNA (referred to as the 5' UTR and 3' UTR). The UTRs are parts of the mRNA that will not be translated into protein, but they have other functions, such as ribosome binding

We can apply the same reasoning to an F1 monohybrid cross. With seed shape in pea plants as the heritable character, the genotype of F1 plants is Rr. Segregation in a heterozygous plant is like flipping a coin in terms of calculating the probability of each outcome: Each egg produced has a 1 ⁄ 2 chance of carrying the dominant allele (R) and a 1 ⁄ 2 chance of carrying the recessive allele (r). The same odds apply to each sperm cell produced.

For a particular F2 plant to have wrinkled seeds, the recessive trait, both the egg and the sperm that come together must carry the r allele. The probability that an r allele will be present in both gametes at fertilization is found by multiplying 1 ⁄ 2 (the probability that the egg will have an r) * 1 ⁄ 2 (the probability that the sperm will have an r). Thus, the multiplication rule tells us that the probability of an F2 plant having wrinkled seeds (rr) is 1 ⁄ 4 (Figure 14.9). Likewise, the probability of an F2 plant carrying both dominant alleles for seed shape (RR) is 1 ⁄ 4.

Frequency of Dominant Alleles: Although you might assume that the dominant allele for a particular character would be more common than the recessive allele, this is not always the case.

For an example of a rare dominant allele, about one baby out of 400 in the United States is born with extra fingers or toes, a condition known as polydactyly. Some cases are caused by the presence of a dominant allele

Additional Evidence That DNA Is the Genetic Material: Further evidence that DNA is the genetic material came from the laboratory of biochemist Erwin Chargaff. DNA was known to be a polymer of nucleotides, each having three components: a nitrogenous (nitrogen-containing) base, a pentose sugar called deoxyribose, and a phosphate group (Figure 16.5). The base can be adenine (A), thymine (T), guanine (G), or cytosine (C). Chargaff analyzed the base composition of DNA from a number of different organisms. In 1950, he reported that the base composition of DNA varies from one species to another.

For example, he found that 32.8% of sea urchin DNA nucleotides have the base A, whereas 30.4% of human DNA nucleotides have the base A and only 24.7% of the DNA nucleotides from the bacterium E. coli have the base A. Chargaff's evidence of molecular diversity among species, which most scientists had presumed to be absent from DNA, made DNA a more credible candidate for the genetic material.

The two chromosomes of a pair have the same length, centromere position, and staining pattern: These are called homologous chromosomes (or homologs). Both chromosomes of each pair carry genes controlling the same inherited characters.

For example, if a gene for eye color is situated at a particular locus on a certain chromosome, then its homologous chromosome (its homolog) will also have a version of the eye-color gene at the equivalent locus.

Ribozymes: The idea of a catalytic role for the RNAs in the spliceosome arose from the discovery of ribozymes, RNA molecules that function as enzymes. In some organisms, RNA splicing can occur without proteins or even additional RNA molecules: The intron RNA functions as a ribozyme and catalyzes its own excision!

For example, in the ciliate protist Tetrahymena, self-splicing occurs in the production of ribosomal RNA (rRNA), a component of the organism's ribosomes. The pre-rRNA actually removes its own introns. The discovery of ribozymes rendered obsolete the idea that all biological catalysts are proteins.

Evidence from Studying Metabolic Defects: In 1902, British physician Archibald Garrod was the first to suggest that genes dictate phenotypes through enzymes, proteins that catalyze specific chemical reactions in the cell. He postulated that the symptoms of an inherited disease reflect an inability to make a particular enzyme. He later referred to such diseases as "inborn errors of metabolism."

For example, people with a disease called alkaptonuria have black urine because it contains a chemical called alkapton, which darkens upon exposure to air. Garrod reasoned that most people have an enzyme that breaks down alkapton, whereas people with alkaptonuria have inherited an inability to make that enzyme, so alkapton is expelled in their urine.

Comparison of Asexual and Sexual Reproduction: Only organisms that reproduce asexually have offspring that are exact genetic copies of themselves. . In asexual reproduction, a single individual (like a yeast cell or an amoeba; see Figure 12.2a) is the sole parent and passes copies of all its genes to its offspring without the fusion of gametes.

For example, single-celled eukaryotic organisms can reproduce asexually by mitotic cell division, in which DNA is copied and allocated equally to two daughter cells. The genomes of the offspring are virtually exact copies of the parent's genome. Some multicellular organisms are also capable of reproducing asexually (Figure 13.2).

The second instance of molecular recognition is the pairing of the tRNA anticodon with the appropriate mRNA codon. If one tRNA variety existed for each mRNA codon specifying an amino acid, there would be 61 tRNAs (see Figure 17.6). In bacteria, however, there are only about 45 tRNAs, signifying that some tRNAs must be able to bind to more than one codon. Such versatility is possible because the rules for base pairing between the third nucleotide base of a codon and the corresponding base of a tRNA anticodon are relaxed compared to those at other codon positions.

For example, the nucleotide base U at the 5' end of a tRNA anticodon can pair with either A or G in the third position (at the 3' end) of an mRNA codon. The flexible base pairing at this codon position is called wobble. Wobble explains why the synonymous codons for a given amino acid most often differ in their third nucleotide base. Accordingly, a tRNA with the anticodon 3'-UCU-5' can base-pair with either the mRNA codon 5'-AGA-3' or 5'-AGG-3', both of which code for arginine (see Figure 17.6).

In addition to the gene for the viral protein that prevents transcription, a few other prophage genes may be expressed during lysogeny. Expression of these genes may alter the host's phenotype, a phenomenon that can have important medical significance.

For example, the three species of bacteria that cause the human diseases diphtheria, botulism, and scarlet fever would not be so harmful to humans without certain prophage genes that cause the host bacteria to make toxins. And the difference between the E. coli strain in our intestines and the O157:H7 strain that has caused several deaths by food poisoning appears to be the presence of toxin genes of prophages in the O157:H7 strain.

Nüsslein-Volhard and Wieschaus began their search for segmentation genes by exposing flies to a mutagenic chemical that affected the flies' gametes. They mated the mutagenized flies and then scanned their descendants for dead embryos or larvae with abnormal segmentation or other defects.

For example, to find genes that might set up the anterior-posterior axis, they looked for embryos or larvae with abnormal ends, such as two heads or two tails, predicting that such abnormalities would arise from mutations in maternal genes required for correctly setting up the offspring's head or tail end.

The Multiplication and Addition Rules Applied to Monohybrid Crosses: How do we determine the probability that two or more independent events will occur together in some specific combination?

For example, what is the chance that two coins tossed simultaneously will both land heads up? The multiplication rule states that to determine this probability, we multiply the probability of one event (one coin coming up heads) by the probability of the other event (the other coin coming up heads). By the multiplication rule, then, the probability that both coins will land heads up is 1 ⁄ 2 * 1 ⁄ 2 = 1 ⁄ 4.

Mendel observed the same pattern of inheritance in six other characters, each represented by two distinctly different traits (Table 14.1).

For example, when Mendel crossed a truebreeding variety that produced smooth, round pea seeds with one that produced wrinkled seeds, all the F1 hybrids produced round seeds; this is the dominant trait for seed shape.

Unlike somatic cells, gametes contain a single set of chromosomes. Such cells are called haploid cells, and each has a haploid number of chromosomes (n).

For humans, the haploid number is 23 (n = 23). The set of 23 consists of the 22 autosomes plus a single sex chromosome. An unfertilized egg contains an X chromosome; a sperm contains either an X or a Y chromosome.

Scientists can now measure the expression of thousands of genes at one time. DNA technology makes such studies possible; with automation, they are easily performed on a large scale. By uncovering gene interactions and providing clues to gene function, DNA microarray assays and RNA-seq may contribute to a better understanding of diseases and suggest new diagnostic techniques or therapies.

For instance, comparing patterns of gene expression in breast cancer tumors and noncancerous breast tissue has already resulted in more informed and effective treatment protocols (see Figure 18.27). Ultimately, information from these methods should provide a grander view of how ensembles of genes interact to form an organism and maintain its vital systems.

As we have seen, emerging viruses are generally not new; rather, they are existing viruses that mutate, disseminate more widely in the current host species, or spread to new host species. Changes in host behavior or environmental changes can increase the viral traffic responsible for emerging diseases.

For instance, new roads built through remote areas can allow viruses to spread between previously isolated human populations. Also, the destruction of forests to expand cropland can bring humans into contact with other animals that may host viruses capable of infecting humans. Finally, genetic mutations and changes in host ranges can allow viruses to jump from one species to another. Many viruses, including chikungunya, mentioned earlier, can be transmitted by mosquitoes. A dramatic expansion of the disease caused by chikungunya occurred in the mid-2000s when a mutation in the virus allowed it to infect not only the mosquito species Aedes aegypti, but also the related Aedes albopictus. Promotion of the use of insecticides and mosquito netting over beds are crucial tools in public health attempts to prevent diseases carried by mosquitoes (Figure 19.11).

A simple example of alternative RNA splicing is shown in Figure 18.13 for the troponin T gene, which encodes two different (though related) proteins. Other genes code for many more possible products.

For instance, researchers have found a Drosophila gene with enough alternatively spliced exons to generate about 19,000 membrane proteins that have different extracellular domains. At least 17,500 (94%) of the alternative mRNAs are actually synthesized. Each developing nerve cell in the fly appears to synthesize a different form of the protein, which acts as unique identifier on the cell surface and helps prevent excessive overlap of nerve cells during development of the nervous system.

First, in many cases, a eukaryotic gene can code for a set of closely related polypeptides via a process called alternative splicing, which you will learn about later in this chapter. Second, quite a few genes code for RNA molecules that have important functions in cells even though they are never translated into protein.

For now, we will focus on genes that do code for polypeptides. (Note that it is common to refer to these gene products as proteins—a practice you will encounter in this book—rather than more precisely as polypeptides.)

Whether human characters are more influenced by genes or the environment—in everyday terms, nature versus nurture—is a debate that we will not attempt to settle here. We can say, however, that a genotype generally is not associated with a rigidly defined phenotype, but rather with a range of phenotypic possibilities due to environmental influences (Figure 14.14).

For some characters, such as the ABO blood group system, the phenotypic range has no breadth whatsoever; that is, a given genotype mandates a very specific phenotype. Other characters, such as a person's blood count of red and white cells, vary quite a bit, depending on such factors as the altitude, the customary level of physical activity, and the presence of infectious agents.

Degrees of Dominance: Alleles can show different degrees of dominance and recessiveness in relation to each other. In Mendel's classic pea crosses, the F1 offspring always looked like one of the two parental varieties because one allele in a pair showed complete dominance over the other. In such situations, the phenotypes of the heterozygote and the dominant homozygote are indistinguishable (see Figure 14.6).

For some genes, however, neither allele is completely dominant, and the F1 hybrids have a phenotype somewhere between those of the two parental varieties. This phenomenon, called incomplete dominance, is seen when red snapdragons are crossed with white snapdragons: All the F1 hybrids have pink flowers (Figure 14.10). This third, intermediate phenotype results from flowers of the heterozygotes having less red pigment than the red homozygotes. (This is unlike the case of Mendel's pea plants, where the Pp heterozygotes make enough pigment for the flowers to be purple, indistinguishable from those of PP plants.)

Recall that the trp repressor protein is inactive by itself and requires tryptophan as a corepressor in order to bind to the operator. The lac repressor, in contrast, is active by itself, binding to the operator and switching the lac operon off. In this case, a specific small molecule, called an inducer, inactivates the repressor.

For the lac operon, the inducer is allolactose, an isomer of lactose formed in small amounts from lactose that enters the cell. In the absence of lactose (and hence allolactose), the lac repressor is in its active shape and binds to the operator; thus, the genes of the lac operon are silenced (Figure 18.4a). If lactose is added to the cell's surroundings, allolactose binds to the lac repressor and alters its shape so the repressor can no longer bind to the operator. Without the lac repressor bound, the lac operon is transcribed into mRNA, and the enzymes for using lactose are made (Figure 18.4b).

Types of Small-Scale Mutations: Let's now consider how small-scale mutations affect proteins. We should first note that many mutations occur in regions outside of protein-coding genes, and any potential effect they have on the phenotype of the organism may be subtle and hard to detect.

For this reason, here we'll concentrate on mutations within protein-coding genes. Small-scale mutations within a gene can be divided into two general categories: (1) single nucleotide-pair substitutions and (2) nucleotide-pair insertions or deletions. Insertions and deletions can involve one or more nucleotide pairs

Mendel worked out the second law of inheritance by following two characters at the same time, such as seed color and seed shape. Seeds (peas) may be either yellow or green. They also may be either round (smooth) or wrinkled.

From singlecharacter crosses, Mendel knew that the allele for yellow seeds is dominant (Y), and the allele for green seeds is recessive (y). For the seed-shape character, the allele for round is dominant (R), and the allele for wrinkled is recessive (r)

Morgan's Choice of Experimental Organism: Many times in the history of biology, important discoveries have come to those insightful or lucky enough to choose an experimental organism suitable for the research problem being tackled. Mendel chose the garden pea because a number of distinct varieties were available. For his work, Morgan selected a species of fruit fly, Drosophila melanogaster, a common insect that feeds on the fungi growing on fruit.

Fruit flies are prolific breeders; a single mating will produce hundreds of offspring, and a new generation can be bred every two weeks. Morgan's laboratory began using this convenient organism for genetic studies in 1907 and soon became known as "the fly room."

Genetic Analysis of Early Development: Scientific Inquiry: Edward B. Lewis was a visionary American biologist who, in the 1940s, first showed the value of the genetic approach to studying embryonic development in Drosophila. Lewis studied bizarre mutant flies with developmental defects that led to extra wings or legs in the wrong place (Figure 18.20). He located the mutations on the fly's genetic map, thus connecting the developmental abnormalities to specific genes. This research supplied the first concrete evidence that genes somehow direct the developmental processes studied by embryologists. The genes Lewis discovered, called homeotic genes, are regulatory genes that control pattern formation in the late embryo, larva, and adult.

Further insight into pattern formation during early embryonic development did not come for another 30 years, when two researchers in Germany, Christiane NüssleinVolhard and Eric Wieschaus, set out to identify all the genes that affect segment formation in Drosophila. The project was daunting for three reasons. The first was the sheer number of Drosophila genes, now known to total about 14,000. The genes affecting segmentation might be just a few needles in a haystack or might be so numerous and varied that the scientists would be unable to make sense of them. Second, mutations affecting a process as fundamental as segmentation would surely be embryonic lethals, mutations with phenotypes causing death at the embryonic or larval stage. Because organisms with embryonic lethal mutations never reproduce, they cannot be bred for study.

The CRISPR-Cas system was discovered during a study of repetitive DNA sequences present in the genomes of many prokaryotes. These sequences, which puzzled scientists, were named clustered regularly interspaced short palindromic repeats (CRISPRs) because each sequence read the same forward and backward (a palindrome), with different stretches of "spacer DNA" in between the repeats. At first, scientists assumed the spacer DNA sequences were random and meaningless, but analysis by several research groups showed that each spacer sequence corresponded to DNA from a particular phage that had infected the cell.

Further studies revealed that particular nuclease proteins interact with the CRISPR region. These nucleases, called Cas (CRISPR-associated) proteins, can identify and cut phage DNA, thereby defending the bacterium against phage infection.

Because Beadle and Tatum set up their experimental conditions so that each mutant was defective in a single gene, the collected results, taken together, provided strong support for a working hypothesis they had proposed earlier. The one gene-one enzyme hypothesis, as they dubbed it, states that the function of a gene is to dictate the production of a specific enzyme.

Further support for this hypothesis came from experiments that identified the specific enzymes lacking in the mutants. Beadle and Tatum shared a Nobel Prize in 1958 for "their discovery that genes act by regulating definite chemical events" (in the words of the Nobel committee).

About half a dozen changes must occur at the DNA level for a cell to become fully cancerous. These changes usually include the appearance of at least one active oncogene and the mutation or loss of several tumor-suppressor genes.

Furthermore, since mutant tumor-suppressor alleles are usually recessive, in most cases mutations must knock out both alleles in a cell's genome to block tumor suppression. (Most oncogenes, on the other hand, behave as dominant alleles.)

Evidence That DNA Can Transform Bacteria: In 1928, a British medical officer named Frederick Griffith was trying to develop a vaccine against pneumonia. He was studying Streptococcus pneumoniae, a bacterium that causes pneumonia in mammals. Griffith had two strains (varieties) of the bacterium, one pathogenic (disease-causing) and one nonpathogenic (harmless). He was surprised to find that when he killed the pathogenic bacteria with heat and then mixed the cell remains with living bacteria of the nonpathogenic strain, some of the living cells became pathogenic (Figure 16.2).

Furthermore, this newly acquired trait of pathogenicity was inherited by all the descendants of the transformed bacteria. Apparently, some chemical component of the dead pathogenic cells caused this heritable change, although the identity of the substance was not known. Griffith called the phenomenon transformation, now defined as a change in genotype and phenotype due to the assimilation of external DNA by a cell. Later work by Oswald Avery, Maclyn McCarty, and Colin MacLeod identified the transforming substance as DNA.

How easily could this happen? In 2011, scientists working with ferrets, small mammals that are animal models for human flu, found out that only a few mutations of the avian flu virus would allow infection of cells in the human nasal cavity and windpipe.

Furthermore, when the scientists transferred nasal swabs serially from ferret to ferret, the virus became transmissible through the air. Reports of this startling discovery at a scientific conference ignited a firestorm of debate about whether to publish the results and led to an ongoing reevaluation of the federal policies governing this type of experiment in the United States. The risks of doing this type of research (what if the new virus escapes or the procedure falls into the hands of bioterrorists?) must be considered in relation to the risks of not doing it—the possibility that we will be unable to combat new, more transmissible viruses because we lack an understanding of how they develop.

Using Restriction Enzymes to Make a Recombinant DNA Plasmid

Gene cloning and genetic engineering generally rely on the use of enzymes that cut DNA molecules at a limited number of specific locations. These enzymes, called restriction endonucleases, or restriction enzymes, were discovered in the late 1960s by biologists doing basic research on bacteria. Restriction enzymes protect the bacterial cell by cutting up foreign DNA from other organisms or phages (see Concept 19.2).

This example illustrates the main point of this chapter: The DNA inherited by an organism leads to specific traits by dictating the synthesis of proteins and of RNA molecules involved in protein synthesis. In other words, proteins are the link between genotype and phenotype.

Gene expression is the process by which DNA directs the synthesis of proteins (or, in some cases, just RNAs). The expression of genes that code for proteins includes two stages: transcription and translation. This chapter describes the flow of information from gene to protein and explains how genetic mutations affect organisms through their proteins. Understanding the processes of gene expression will allow us to revisit the concept of the gene in more detail at the end of the chapter.

Although more elaborate techniques were required to decode mixed triplets such as AUA and CGA, all 64 codons were deciphered by the mid-1960s. As Figure 17.6 shows, 61 of the 64 triplets code for amino acids. The three codons that do not designate amino acids are "stop" signals, or termination codons, marking the end of translation. Notice that the codon AUG has a dual function: It codes for the amino acid methionine (Met, or M) and also functions as a "start" signal, or initiation codon.

Genetic messages usually begin with the mRNA codon AUG, which signals the proteinsynthesizing machinery to begin translating the mRNA at that location. (Because AUG also stands for methionine, polypeptide chains begin with methionine when they are synthesized. However, an enzyme may subsequently remove this starter amino acid from the chain.)

Genetics

Genetics is the scientific study of heredity and inherited variation. In this unit, you'll learn about genetics at multiple levels, from organisms to cells to molecules. We begin by examining how chromosomes pass from parents to offspring in sexually reproducing organisms. The processes of meiosis (a special type of cell division) and fertilization (the fusion of sperm and egg, as seen in the small photo) maintain a species' chromosome count during the sexual life cycle. We will describe the cellular mechanics of meiosis and explain how this process differs from mitosis. Finally, we will consider how both meiosis and fertilization contribute to genetic variation, such as that seen in Figure 13.1.

A recent study may underscore the protective role of p53 while illuminating a long-standing research question: Why is cancer so rare among elephants? The incidence of cancer among elephants in zoo-based studies has been estimated at about 3%, compared to closer to 30% for humans.

Genome sequencing revealed that elephants have 20 copies of the p53 gene, compared to one copy in humans, other mammals, and even manatees, elephants' closest living relatives. There are undoubtedly other underlying reasons, but the correlation between low cancer rate and extra copies of the p53 gene bears further investigation.

Studying the Expression of Interacting Groups of Genes: A major goal of biologists is to learn how genes act together to produce and maintain a functioning organism. Now that the genomes of a number of species have been sequenced, it is possible to study the expression of large groups of genes—the so-called systems approach. Researchers use what is known about the whole genome to investigate which genes are transcribed in different tissues or at different stages of development. One aim is to identify networks of gene expression across an entire genome.

Genome-wide expression studies can be carried out using DNA microarray assays. A DNA microarray consists of tiny amounts of a large number of single-stranded DNA fragments representing different genes fixed to a glass slide in a tightly spaced array, or grid, of dots. (The microarray is also called a DNA chip by analogy to a computer chip.) Ideally, these fragments represent all the genes of an organism. The mRNAs from cells under study are reverse-transcribed into cDNAs (see Figure 20.10), and a fluorescent label is added so the cDNAs can be used as probes on the microarray. Different fluorescent labels are used for different cell samples so that multiple samples can be tested in the same experiment. The resulting pattern of colored dots, shown in an actual-size microarray in Figure 20.12, reveals the dots to which each probe was bound and thus the genes that are expressed in the cell samples being tested. Microarray technology started taking off after several papers about it were published in 1995 since then more sophisticated applications have been developed and are in use.

The blocking of gene expression by siRNAs is referred to as RNA interference (RNAi), and it is used in the laboratory as a means of disabling specific genes to investigate their function. How did the RNAi pathway evolve? As you will learn in Concept 19.2, some viruses have double-stranded RNA genomes.

Given that the cellular RNAi pathway can process double-stranded RNAs into homing devices that lead to destruction of related RNAs, some scientists think that this pathway may have evolved as a natural defense against infection by such viruses.

The RNA animal viruses with the most complicated replicative cycles are the retroviruses (class VI). These viruses have an enzyme called reverse transcriptase that transcribes an RNA template into DNA, an RNA S DNA information flow that is the opposite of the usual direction. This unusual phenomenon is the source of the name retroviruses (retro means "backward"). Of particular medical importance is HIV (human immunodeficiency virus), the retrovirus shown in Figure 19.1 that causes AIDS (acquired immunodeficiency syndrome).

HIV and other retroviruses are enveloped viruses that contain two identical molecules of single-stranded RNA and two molecules of reverse transcriptase.

Prion aggregation interferes with normal cellular functions and causes disease symptoms. This model was greeted with much skepticism when it was first proposed by Stanley Prusiner in the early 1980s, but it is now widely accepted. Prusiner was awarded the Nobel Prize in 1997 for his work on prions.

He has recently proposed that prions are also involved in neurodegenerative diseases such as Alzheimer's and Parkinson's disease. There are many outstanding questions about these small infectious agents.

Multifactorial Disorders: The hereditary diseases we have discussed so far are sometimes described as simple Mendelian disorders because they result from abnormality of one or both alleles at a single genetic locus. Many more people are susceptible to diseases that have a multifactorial basis—a genetic component plus a significant environmental influence.

Heart disease, diabetes, cancer, alcoholism, certain mental illnesses such as schizophrenia and bipolar disorder, and many other diseases are multifactorial. In these cases, the hereditary component is polygenic. For example, many genes affect cardiovascular health, making some of us more prone than others to heart attacks and strokes. No matter what our genotype, however, our lifestyle has a tremendous effect on phenotype for cardiovascular health and other multifactorial characters. Exercise, a healthful diet, abstinence from smoking, and an ability to handle stressful situations all reduce our risk of heart disease and some types of cancer.

Somehow, T2 could reprogram its host cell to produce viruses. But which viral component—protein or DNA—was responsible?

Hershey and Chase answered this question by devising an experiment showing that only one of the two components of T2 actually enters the E. coli cell during infection (Figure 16.4).

Dominantly Inherited Disorders: Although many harmful alleles are recessive, a number of human disorders are due to dominant alleles. One example is achondroplasia, a form of dwarfism that occurs in one of every 25,000 people.

Heterozygous individuals have the dwarf phenotype (Figure 14.18). Therefore, all people who do not have achondroplasia—99.99% of the population—are homozygous for the recessive allele. Like the presence of extra fingers or toes mentioned earlier, achondroplasia is a trait for which the recessive allele is much more prevalent than the corresponding dominant allele.

Cracking the Code: Molecular biologists cracked the genetic code of life in the early 1960s when a series of elegant experiments disclosed the amino acid translations of each of the RNA codons. The first codon was deciphered in 1961 by Marshall Nirenberg, of the National Institutes of Health, along with his colleagues. Nirenberg synthesized an artificial mRNA by linking together many identical RNA nucleotides containing uracil as their base. No matter where the genetic message started or stopped, it could contain only one codon (UUU) over and over. Nirenberg added this "poly-U" polynucleotide to a test-tube mixture containing amino acids, ribosomes, and the other components required for protein synthesis.

His artificial system translated the poly-U mRNA into a polypeptide containing many units of the amino acid phenylalanine (Phe, or F), strung together as a long polyphenylalanine chain. Thus, Nirenberg determined that the mRNA codon UUU specifies the amino acid phenylalanine. Soon, the amino acids specified by the codons AAA, GGG, and CCC were also identified.

Sturtevant hypothesized that the percentage of recombinant offspring, the recombination frequency, calculated from experiments like the one in Figures 15.9 and 15.10, depends on the distance between genes on a chromosome. He assumed that crossing over is a random event, with the chance of crossing over approximately equal at all points along a chromosome. Based on these assumptions, Sturtevant predicted that the farther apart two genes are, the higher the probability that a crossover will occur between them and therefore the higher the recombination frequency.

His reasoning was simple: The greater the distance between two genes, the more points there are between them where crossing over can occur. Using recombination data from various fruit fly crosses, Sturtevant proceeded to assign relative positions to genes on the same chromosomes—that is, to map genes.

In fact, the pathogen replicated only within the host it infected. In further experiments, Beijerinck showed that unlike bacteria used in the lab at that time, the mysterious agent of mosaic disease could not be cultivated on nutrient media in test tubes or petri dishes. Beijerinck imagined a replicating particle much smaller and simpler than a bacterium, and he is generally credited with being the first scientist to voice the concept of a virus.

His suspicions were confirmed in 1935 when the American scientist Wendell Stanley crystallized the infectious particle, now known as tobacco mosaic virus (TMV). Subsequently, TMV and many other viruses were actually seen with the help of the electron microscope.

Meiosis I

Homologous chromosomes separate. Haploid cells with duplicated chromosomes

In contrast, the two chromosomes of a homologous pair are individual chromosomes that were inherited from each parent.

Homologs appear alike in the microscope, but they may have different versions of genes at corresponding loci; each version is called an allele of that gene (see Figure 14.4). Homologs are not associated with each other in any obvious way except during meiosis. Figure 13.8 describes in detail the stages of the two divisions of meiosis for an animal cell whose diploid number is 6. Study this figure thoroughly before going on.

Genetic variation produced in sexual life cycles contributes to evolution:

How do we account for the genetic variation of the family members in Figure 13.1? As you will learn more about in later chapters, mutations are the original source of genetic diversity. These changes in an organism's DNA create the different versions of genes, known as alleles. Once these differences arise, reshuffling of the alleles during sexual reproduction produces the variation that results in each member of a sexually reproducing population having a unique combination of traits.

Viral Diseases in Animals: A viral infection can produce symptoms by a number of different routes. Viruses may damage or kill cells by causing the release of hydrolytic enzymes from lysosomes. Some viruses cause infected cells to produce toxins that lead to disease symptoms, and some have molecular components that are toxic, such as envelope proteins.

How much damage a virus causes depends partly on the ability of the infected tissue to regenerate by cell division. People usually recover completely from colds because the epithelium of the respiratory tract, which the viruses infect, can efficiently repair itself. In contrast, damage inflicted by poliovirus to mature nerve cells is permanent because these cells do not divide and usually cannot be replaced. Many of the temporary symptoms associated with viral infections, such as fever and body aches, actually result from the body's own efforts to defend itself against infection rather than from cell death caused by the virus.

Today, we know that genes—Mendel's "factors"—are segments of DNA located along chromosomes. We can see the location of a particular gene by tagging chromosomes with a fluorescent dye that highlights that gene. For example, the four yellow dots in Figure 15.1 mark a specific gene on a pair of homologous human chromosomes. (The chromosomes have duplicated, so each chromosome has two copies of the allele, one on each sister chromatid.)

However, Gregor Mendel's "hereditary factors" were purely an abstract concept when he proposed their existence in 1860. At that time, no cellular structures had been identified that could house these imaginary units, and most biologists were skeptical about Mendel's proposed laws of inheritance.

Recall that a female fly has two X chromosomes (XX), while a male fly has an X and a Y (XY). The correlation between the trait of white eye color and the male sex of the affected F2 flies suggested to Morgan that the gene involved in his white-eyed mutant was located exclusively on the X chromosome, with no corresponding allele present on the Y chromosome. His reasoning can be followed in Figure 15.4. For a male, a single copy of the mutant allele would confer white eyes; since a male has only one X chromosome, there can be no wild-type allele (w+ ) present to mask the recessive allele.

However, a female could have white eyes only if both her X chromosomes carried the recessive mutant allele (w). This was impossible for the F2 females in Morgan's experiment because all the F1 fathers had red eyes, so each F2 female received a w+ allele on the X chromosome inherited from her father.

Noncoding RNAs play multiple roles in controlling gene expression: Genome sequencing has revealed that protein-coding DNA accounts for only 1.5% of the human genome and a similarly small percentage of the genomes of many other multicellular eukaryotes. A very small fraction of the non-protein-coding DNA consists of genes for RNAs such as ribosomal RNA and transfer RNA. Until recently, scientists assumed that most of the remaining DNA was not transcribed, thinking that since it didn't specify proteins or the few known types of RNA, such DNA didn't contain meaningful genetic information—in fact, it was called "junk DNA."

However, a flood of recent data has contradicted this idea. For example, a massive study of the entire human genome showed that roughly 75% of the genome is transcribed at some point in any given cell. Introns account for only a fraction of this transcribed, nontranslated RNA. These and other results suggest that a significant amount of the genome may be transcribed into non-protein-coding RNAs—also called noncoding RNAs, or ncRNAs—including a variety of small RNAs. Researchers are uncovering more evidence of the biological roles of these ncRNAs every day.

If an X-linked trait is due to a recessive allele, a female will express the phenotype only if she is homozygous for that allele. Because males have only one locus, the terms homozygous and heterozygous lack meaning for describing their X-linked genes; the term hemizygous is used in such cases. Any male receiving the recessive allele from his mother will express the trait. For this reason, far more males than females have X-linked recessive disorders.

However, even though the chance of a female inheriting a double dose of the mutant allele is much less than the probability of a male inheriting a single dose, there are females with X-linked disorders. For instance, color blindness is almost always inherited as an X-linked trait. A color-blind daughter may be born to a color-blind father whose mate is a carrier (see Figure 15.7c). Because the X-linked allele for color blindness is relatively rare, though, the probability that such a man and woman will mate is low.

During transcription, the gene determines the sequence of nucleotide bases along the length of the RNA molecule that is being synthesized. For each gene, only one of the two DNA strands is transcribed. This strand is called the template strand because it provides the pattern, or template, for the sequence of nucleotides in an RNA transcript. For any given gene, the same strand is used as the template every time the gene is transcribed.

However, farther along on the same chromosomal DNA molecule, the opposite strand may function as the template for a different gene. The strand that is used as the template is determined by the orientation of the enzyme that transcribes genes, which in turn depends on the particular DNA sequences associated with that gene.

What exactly is a genomic imprint? It turns out that imprinting can involve either silencing an allele in one type of gamete (egg or sperm) or activating it in the other. In many cases, the imprint seems to consist of methyl (—CH3) groups that are added to cytosine nucleotides of one of the alleles. Such methylation may silence the allele, an effect consistent with evidence that heavily methylated genes are usually inactive (see Concept 18.2).

However, for a few genes, methylation has been shown to activate expression of the allele. This is the case for the Igf2 gene: Methylation of certain cytosines on the paternal chromosome leads to expression of the paternal Igf2 allele, by an indirect mechanism involving chromatin structure and protein-DNA interactions

Pedigrees are a more serious matter when the alleles in question cause disabling or deadly diseases instead of innocuous human variations such as hairline or inability to taste an innocuous chemical.

However, for disorders inherited as simple Mendelian traits, the same techniques of pedigree analysis apply

The Products of Gene Expression: A Developing Story As researchers learned more about proteins, they made revisions to the one gene-one enzyme hypothesis. First of all, not all proteins are enzymes. Keratin, the structural protein of animal hair, and the hormone insulin are two examples of nonenzyme proteins. Because proteins that are not enzymes are nevertheless gene products, molecular biologists began to think in terms of one gene-one protein.

However, many proteins are constructed from two or more different polypeptide chains, and each polypeptide is specified by its own gene. For example, hemoglobin—the oxygen-transporting protein of vertebrate red blood cells— contains two kinds of polypeptides (see Figure 5.18), and thus two genes code for this protein, one for each type of polypeptide. Beadle and Tatum's idea was therefore restated as the one gene-one polypeptide hypothesis. Even this description is not entirely accurate, though.

Human Disorders Due to Chromosomal Alterations: Alterations of chromosome number and structure are associated with a number of serious human disorders. As described earlier, nondisjunction in meiosis results in aneuploidy in gametes and any resulting zygotes. Although the frequency of aneuploid zygotes may be quite high in humans, most of these chromosomal alterations are so disastrous to development that the affected embryos are spontaneously aborted long before birth.

However, some types of aneuploidy appear to upset the genetic balance less than others, so individuals with certain aneuploid conditions can survive to birth and beyond. These individuals have a set of traits—a syndrome— characteristic of the type of aneuploidy. Genetic disorders caused by aneuploidy can be diagnosed before birth by fetal testing (see Figure 14.19).

Although vaccines can prevent some viral illnesses, medical care can do little, at present, to cure most viral infections once they occur. The antibiotics that help us recover from bacterial infections are powerless against viruses. Antibiotics kill bacteria by inhibiting enzymes specific to bacteria but have no effect on eukaryotic or virally encoded enzymes.

However, the few enzymes that are encoded only by viruses have provided targets for other drugs. Most antiviral drugs resemble nucleosides and thus interfere with viral nucleic acid synthesis. One such drug is acyclovir, which impedes herpesvirus replication by inhibiting the viral polymerase that synthesizes viral DNA but not the eukaryotic one. Similarly, azidothymidine (AZT) curbs HIV replication by interfering with the synthesis of DNA by reverse transcriptase.

Incorrectly paired or altered nucleotides can also arise after replication. In fact, maintenance of the genetic information encoded in DNA requires frequent repair of various kinds of damage to existing DNA. DNA molecules are constantly subjected to potentially harmful chemical and physical agents, such as X-rays, as we'll discuss in Concept 17.5. In addition, DNA bases may undergo spontaneous chemical changes under normal cellular conditions.

However, these changes in DNA are usually corrected before they become permanent changes—mutations—perpetuated through successive replications. Each cell continuously monitors and repairs its genetic material. Because repair of damaged DNA is so important to the survival of an organism, it is no surprise that many different DNA repair enzymes have evolved. Almost 100 are known in E. coli, and about 170 have been identified so far in humans

Meiosis reduces the number of chromosome sets from diploid to haploid: Several steps of meiosis closely resemble corresponding steps in mitosis. Meiosis, like mitosis, is preceded by the duplication of chromosomes.

However, this single duplication is followed by not one but two consecutive cell divisions, called meiosis I and meiosis II. These two divisions result in four daughter cells (rather than the two daughter cells of mitosis), each with only half as many chromosomes as the parent cell—one set, rather than two.

Are viruses living or nonliving? Early on, they were considered biological chemicals; the Latin root for virus means "poison." Viruses are capable of causing a wide variety of diseases, so researchers in the late 1800s saw a parallel with bacteria and proposed that viruses were the simplest of living forms.

However, viruses cannot reproduce or carry out metabolic activities outside of a host cell. Most biologists studying viruses today would probably agree that they are not alive but exist in a shady area between life-forms and chemicals. The simple phrase used recently by two researchers describes them aptly enough: Viruses lead "a kind of borrowed life."

Abnormal Chromosome Number:

Ideally, the meiotic spindle distributes chromosomes to daughter cells without error. But there is an occasional mishap, called a nondisjunction, in which the members of a pair of homologous chromosomes do not move apart properly during meiosis I or sister chromatids fail to separate during meiosis II (Figure 15.13). In nondisjunction, one gamete receives two of the same type of chromosome and another gamete receives no copy. The other chromosomes are usually distributed normally

Cancer cells are frequently found to contain chromosomes that have broken and rejoined incorrectly, translocating fragments from one chromosome to another (see Figure 15.14). Having learned how gene expression is regulated, you can now see the possible consequences of such translocations.

If a translocated proto-oncogene ends up near an especially active promoter (or other control element), its transcription may increase, making it an oncogene. The second main type of genetic change, amplification, increases the number of copies of the proto-oncogene in the cell through repeated gene duplication (discussed in Concept 21.5). The third possibility is a point mutation either in the promoter or an enhancer that controls a proto-oncogene, causing an increase in its expression, or in the coding sequence of the proto-oncogene, changing the gene's product to a protein that is more active or more resistant to degradation than the normal protein. These mechanisms can lead to abnormal stimulation of the cell cycle and put the cell on the path to becoming a cancer cell.

The Testcross: Given a purple-flowered pea plant, we cannot tell if it is homozygous (PP) or heterozygous (Pp) because both genotypes result in the same purple phenotype. To determine the genotype, we can cross this plant with a white-flowered plant (pp), which will make only gametes with the recessive allele (p). The allele in the gamete contributed by the purple-flowered plant of unknown genotype will therefore determine the appearance of the offspring (Figure 14.7)

If all the offspring of the cross have purple flowers, then the purple-flowered mystery plant must be homozygous for the dominant allele, because a PP * pp cross produces all Pp offspring. But if both the purple and the white phenotypes appear among the offspring, then the purple-flowered parent must be heterozygous. The offspring of a Pp * pp cross will be expected to have a 1:1 phenotypic ratio. Breeding an organism of unknown genotype with a recessive homozygote is called a testcross because it can reveal the genotype of that organism. The testcross was devised by Mendel and continues to be used by geneticists.

The disaccharide lactose (milk sugar) is available to E. coli in the human colon if the host drinks milk or eats a dairy product. Lactose metabolism begins with hydrolysis of the disaccharide into its component monosaccharides (glucose and galactose), a reaction catalyzed by the enzyme β-galactosidase. Only a few molecules of this enzyme are present in an E. coli cell growing in the absence of lactose.

If lactose is added to the bacterium's environment, however, the number of β-galactosidase molecules in the cell increases 1,000-fold within about 15 minutes. How can a cell ramp up enzyme production this quickly? The gene for β-galactosidase (lacZ) is part of the lac operon which includes two other genes coding for enzymes that function in the use of lactose (Figure 18.4). The entire transcription unit is under the command of one main operator and promoter. The regulatory gene, lacI, located outside the lac operon, codes for an allosteric repressor protein that can switch off the lac operon by binding to the lac operator. So far, this sounds just like regulation of the trp operon, but there is one important difference.

The alternative hypothesis is that the two pairs of alleles segregate independently of each other. In other words, genes are packaged into gametes in all possible allelic combinations, as long as each gamete has one allele for each gene (see Figure 13.11). In our example, an F1 plant will produce four classes of gametes in equal quantities: YR, Yr, yR, and yr.

If sperm of the four classes fertilize eggs of the four classes, there will be 16 (4 * 4) equally probable ways in which the alleles can combine in the F2 generation, as shown on the right side of Figure 14.8. These combinations result in four phenotypic categories with a ratio of 9:3:3:1 (nine yellow round to three green round to three yellow wrinkled to one green wrinkled): When Mendel did the experiment and classified the F2 offspring, his results were close to the predicted 9:3:3:1 phenotypic ratio, supporting the hypothesis that the alleles for one gene—controlling seed color, for example—segregate into gametes independently of the alleles of any other gene, such as seed shape.

Tryptophan functions in this system as a corepressor, a small molecule that cooperates with a repressor protein to switch an operon off. As tryptophan accumulates, more tryptophan molecules associate with trp repressor molecules, which can then bind to the trp operator and shut down production of the tryptophan pathway enzymes.

If the cell's tryptophan level drops, many fewer trp repressor proteins would have tryptophan bound, rendering them inactive; they would dissociate from the operator, allowing transcription of the operon's genes to resume. The trp operon is one example of how gene expression can respond to changes in the cell's internal and external environment.

But what about cells whose genome must persist virtually unchanged from an organism to its offspring over many generations?

If the chromosomes of germ cells became shorter in every cell cycle, essential genes would eventually be missing from the gametes they produce. However, this does not occur: An enzyme called telomerase catalyzes the lengthening of telomeres in eukaryotic germ cells, thus restoring their original length and compensating for the shortening that occurs during DNA replication. This enzyme contains its own RNA molecule that it uses as a template to artificially "extend" the leading strand, allowing the lagging strand to maintain a given length. Telomerase is not active in most human somatic cells, but its activity varies from tissue to tissue. The activity of telomerase in germ cells results in telomeres of maximum length in the zygote.

Consider, for instance, an individual Escherichia coli (E. coli) cell living in the erratic environment of a human colon, dependent for its nutrients on the whimsical eating habits of its host. If the environment is lacking in the amino acid tryptophan, which the bacterium needs to survive, the cell responds by activating a metabolic pathway that makes tryptophan from another compound.

If the human host later eats a tryptophanrich meal, the bacterial cell stops producing tryptophan, thus avoiding wasting resources to produce a substance that is readily available from the surrounding solution.

Useful Genetic Vocabulary: An organism that has a pair of identical alleles for a gene encoding a character is called a homozygote and is said to be homozygous for that gene. In the parental generation in Figure 14.5, the purple-flowered pea plant is homozygous for the dominant allele (PP), while the white plant is homozygous for the recessive allele (pp). Homozygous plants "breed true" because all of their gametes contain the same allele—either P or p in this example.

If we cross dominant homozygotes with recessive homozygotes, every offspring will have two different alleles—Pp in the case of the F1 hybrids of our flowercolor experiment (see Figure 14.5). An organism that has two different alleles for a gene is called a heterozygote and is said to be heterozygous for that gene. Unlike homozygotes, heterozygotes produce gametes with different alleles, so they are not true-breeding. For example, P- and p-containing gametes are both produced by our F1 hybrids. Self-pollination of the F1 hybrids thus produces both purple-flowered and whiteflowered offspring.

Now let's see how we can combine the multiplication and addition rules to solve even more complex problems in Mendelian genetics.

Imagine a cross of two pea varieties in which we track the inheritance of three characters. Let's cross a trihybrid with purple flowers and yellow, round seeds (heterozygous for all three genes) with a plant with purple flowers and green, wrinkled seeds (heterozygous for flower color but homozygous recessive for the other two characters).

The only cells of the human body not produced by mitosis are the gametes, which develop from specialized cells called germ cells in the gonads—ovaries in females and testes in males (see Figure 13.5).

Imagine what would happen if human gametes were made by mitosis: They would be diploid like the somatic cells. At the next round of fertilization, when two gametes fused, the normal chromosome number of 46 would double to 92, and each subsequent generation would double the number of chromosomes yet again. This does not happen, however, because in sexually reproducing organisms, gamete formation involves a type of cell division called meiosis. This type of cell division reduces the number of sets of chromosomes from two to one in the gametes, counterbalancing the doubling that occurs at fertilization.

Breast cancer is the second most common form of cancer in the United States, and the first among women. Each year, this cancer strikes over 230,000 women (and some men) in the United States and kills 40,000 (450,000 worldwide). A major problem with understanding breast cancer is its heterogeneity: Tumors differ in significant ways. Identifying differences between types of breast cancer is expected to improve treatment and decrease the mortality rate.

In 2012, The Cancer Genome Atlas Network, sponsored by the National Institutes of Health, published the results of a multi-team effort that used a genomics approach to profile subtypes of breast cancer based on their molecular signatures. Four major types of breast cancer were identified (Figure 18.27). It is now routine to screen for the presence of particular signaling receptors in any breast cancer tumors, and individuals with breast cancer, along with their physicians, can now make more informed decisions about their treatments.

The low frequency of polydactyly indicates that the recessive allele, which results in five digits per appendage, is far more prevalent than the dominant allele in the population.

In Concept 23.3, you will learn how relative frequencies of alleles in a population are affected by natural selection.

In making a primary transcript from a gene, RNA polymerase II transcribes both introns and exons from the DNA, but the mRNA molecule that enters the cytoplasm is an abridged version.

In RNA splicing, the introns are cut out from the molecule and the exons joined together, forming an mRNA molecule with a continuous coding sequence.

Normal Breast Cells in a Milk Duct

In a normal breast cell, the three signal receptors are at normal levels (indicated by +): • ERα+ • PR+ • HER2+

The Basic Principle: Base Pairing to a Template Strand

In a second paper, Watson and Crick stated their hypothesis for how DNA replicates: "Now our model for deoxyribonucleic acid is, in effect, a pair of templates, each of which is complementary to the other. We imagine that prior to duplication the hydrogen bonds are broken, and the two chains unwind and separate. Each chain then acts as a template for the formation on to itself of a new companion chain, so that eventually we shall have two pairs of chains, where we only had one before. Moreover, the sequence of the pairs of bases will have been duplicated exactly." † J. D. Watson and F. H. C. Crick, Genetical implications of the structure of deoxyribonucleic acid, Nature 171:964-967 (1953)

As the environment changes, the population may survive if, in each generation, at least some of its members can cope effectively with the new conditions. Mutations are the original source of different alleles, which are then mixed and matched during meiosis. New and different combinations of alleles may work better than those that previously prevailed.

In a stable environment, though, sexual reproduction seems as if it would be less advantageous than asexual reproduction, which ensures perpetuation of successful combinations of alleles.

Chromatin Remodeling and Effects on Transcription by ncRNAs:

In addition to regulating mRNAs, some ncRNAs act to bring about remodeling of chromatin structure. One example occurs during formation of heterochromatin at the centromere, as studied in a species of yeast.

Morgan's finding of the correlation between a particular trait and an individual's sex provided support for the chromosome theory of inheritance: namely, that a specific gene is carried on a specific chromosome (in this case, an eye color gene on the X chromosome).

In addition, Morgan's work indicated that genes located on a sex chromosome exhibit unique inheritance patterns, which we will discuss in the next section. Recognizing the importance of Morgan's early work, many bright students were attracted to his fly room.

The transmission of hereditary traits has its molecular basis in the replication of DNA, which produces copies of genes that can be passed from parents to offspring.

In animals and plants, reproductive cells called gametes are the vehicles that transmit genes from one generation to the next. During fertilization, male and female gametes (sperm and eggs) unite, passing on genes of both parents to their offspring.

Like mRNA and other types of cellular RNA, transfer RNA molecules are transcribed from DNA templates. In a eukaryotic cell, tRNA, like mRNA, is made in the nucleus and then travels to the cytoplasm, where it will participate in the process of translation.

In both bacterial and eukaryotic cells, each tRNA molecule is used repeatedly, picking up its designated amino acid in the cytosol, depositing this cargo onto a polypeptide chain at the ribosome, and then leaving the ribosome, ready to pick up another of the same amino acid.

The life span of mRNA molecules in the cytoplasm is important in determining the pattern of protein synthesis in a cell. Bacterial mRNA molecules typically are degraded by enzymes within a few minutes of their synthesis. This short life span of mRNAs is one reason bacteria can change their patterns of protein synthesis so quickly in response to environmental changes.

In contrast, mRNAs in multicellular eukaryotes typically survive for hours, days, or even weeks. For instance, the mRNAs for the hemoglobin polypeptides (α-globin and β-globin) in developing red blood cells are unusually stable, and these long-lived mRNAs are translated repeatedly in red blood cells.

Translation is simple in principle but complex in its biochemistry and mechanics, especially in the eukaryotic cell.

In dissecting translation, we'll focus on the slightly less complicated version of the process that occurs in bacteria. We'll first look at the major players in this process.

Genomic imprinting occurs during gamete formation and results in the silencing of a particular allele of certain genes. Because these genes are imprinted differently in sperm and eggs, the offspring expresses only one allele of an imprinted gene, the one that has been inherited from a specific parent— either the female parent or the male parent, depending on the particular gene. The imprints are then transmitted to all body cells during development.

In each generation, the old imprints are "erased" in gamete-producing cells, and the chromosomes of the developing gametes are newly imprinted according to the sex of the individual forming the gametes. In a given species, the imprinted genes are always imprinted in the same way. For instance, a gene imprinted for maternal allele expression is always imprinted this way, generation after generation.

Ribosome Association and Initiation of Translation:

In either bacteria or eukaryotes, the start codon (AUG) signals the start of translation; this is important because it establishes the codon reading frame for the mRNA. In the first step of translation, a small ribosomal subunit binds to both the mRNA and a specific initiator tRNA, which carries the amino acid methionine. . In bacteria, the small subunit can bind the two in either order; it binds the mRNA at a specific RNA sequence, just upstream of the AUG start codon. In the Scientific Skills Exercise, you can work with DNA sequences encoding the ribosomal binding sites on the mRNAs of a group of Eschericia coli genes. In eukaryotes, the small subunit, Figure 17.19 The initiation of translation. Animation: Initiation of Translation with the initiator tRNA already bound, binds to the 5¿ cap of the mRNA and then moves, or scans, downstream along the mRNA until it reaches the start codon; the initiator tRNA then hydrogen-bonds to the AUG start codon.

The Role of Viruses in Cancer: The study of genes associated with cancer, inherited or not, increases our basic understanding of how disruption of normal gene regulation can result in this disease. In addition to the mutations and other genetic alterations described in this section, a number of tumor viruses can cause cancer in various animals, including humans.

In fact, one of the earliest breakthroughs in understanding cancer came in 1911, when Peyton Rous, an American pathologist, discovered a virus that causes cancer in chickens. The Epstein-Barr virus, which causes infectious mononucleosis, has been linked to several types of cancer in humans, notably Burkitt's lymphoma. Papillomaviruses are associated with cancer of the cervix, and a virus called HTLV-1 causes a type of adult leukemia. Viruses play a role in about 15% of the cases of human cancer.

Crossing Over: As a consequence of the independent assortment of chromosomes during meiosis, each of us produces a collection of gametes differing greatly in their combinations of the chromosomes we inherited from our two parents. Figure 13.11 suggests that each chromosome in a gamete is exclusively maternal or paternal in origin.

In fact, this is not the case, because crossing over produces recombinant chromosomes, individual chromosomes that carry genes (DNA) from two different parents (Figure 13.12). In meiosis in humans, an average of one to three crossover events occurs per chromosome pair, depending on the size of the chromosomes and the position of their centromeres.

How might a proto-oncogene—a gene that has an essential function in normal cells—become an oncogene, a cancer causing gene?

In general, an oncogene arises from a genetic change that leads to an increase either in the amount of the proto-oncogene's protein product or in the intrinsic activity of each protein molecule. The genetic changes that convert proto-oncogenes to oncogenes fall into three main categories: movement of DNA within the genome, amplification of a proto-oncogene, and point mutations in a control element or in the proto-oncogene itself (Figure 18.23).

What about the evolutionary significance of small eukaryotic ncRNAs?

In general, extra levels of gene regulation might allow evolution of a higher degree of complexity of form. The versatility of miRNA regulation has therefore led some biologists to hypothesize that an increase in the number of different miRNAs specified by the genome of a given species has allowed morphological complexity to increase over evolutionary time. While this hypothesis is still being evaluated, it is logical to expand the discussion to include all small ncRNAs. Exciting new techniques for rapidly sequencing genomes have allowed biologists to begin asking how many genes for ncRNAs are present in the genome of any given species. A survey of different species supports the notion that siRNAs evolved first, followed by miRNAs and later piRNAs, which are found only in animals. And while there are hundreds of types of miRNAs, there appear to be 60,000 or so types of piRNAs, allowing the potential for very sophisticated gene regulation by piRNAs.

A third source of new viral diseases in humans is the spread of existing viruses from other animals. Scientists estimate that about three-quarters of new human diseases originate in this way. Animals that harbor and can transmit a particular virus but are generally unaffected by it are said to act as a natural reservoir for that virus. For example, the H1N1 virus that caused the 2009 flu pandemic mentioned earlier was likely passed to humans from pigs; for this reason, the disease it caused was originally called "swine flu."

In general, flu epidemics provide an instructive example of the effects of viruses moving between species. There are three types of influenza virus: types B and C, which infect only humans and have never caused an epidemic, and type A, which infects a wide range of animals, including birds, pigs, horses, and humans. Influenza A strains have caused four major flu epidemics among humans in the last 100 years. The worst was the first one, the "Spanish flu" pandemic of 1918-1919, which killed 40-50 million people, including many World War I soldiers

Polyploidy is fairly common in the plant kingdom. The spontaneous origin of polyploid individuals plays an important role in plant evolution (see Concept 24.2). Many species we eat are polyploid: Bananas are triploid, wheat hexaploid (6n), and strawberries octoploid (8n). Polyploid animal species are much less common, but there are a few fishes and amphibians known to be polyploid.

In general, polyploids are more nearly normal in appearance than aneuploids. One extra (or missing) chromosome apparently disrupts genetic balance more than does an entire extra set of chromosomes.

Sets of Chromosomes in Human Cells:

In humans, each somatic cell has 46 chromosomes. During mitosis, the chromosomes become condensed enough to be visible under a light microscope. At this point, they can be distinguished from one another by their size, the position of their centromeres, and the pattern of colored bands produced by certain chromatin-binding stains

Other Methods for Studying Gene Function: Another method for silencing expression of selected genes doesn't alter the genome; instead, it exploits the phenomenon of RNA interference (RNAi), described in Concept 18.3. This experimental approach uses synthetic doublestranded RNA molecules matching the sequence of a particular gene to trigger breakdown of the gene's messenger RNA or to block its translation. In organisms such as the nematode and the fruit fly, RNAi has already proved valuable for analyzing the functions of genes on a large scale. This method is quicker than using the CRISPR-Cas9 system, but it only leads to a temporary reduction of gene expression rather than a permanent gene knockout or alteration.

In humans, ethical considerations prohibit knocking out genes to determine their functions. An alternative approach is to analyze the genomes of large numbers of people with a certain phenotypic condition or disease, such as heart disease or diabetes, to try to find differences they all share compared with people without that condition. The assumption is that these differences may be associated with one or more malfunctioning genes, thus in a sense being naturally occurring gene knockouts. In these large-scale analyses, called genome-wide association studies, researchers look for genetic markers, DNA sequences that vary in the population. In a gene, such sequence variation is the basis of different alleles, as we have seen for sickle-cell disease (see Figure 17.26). And just like the coding sequences of genes, noncoding DNA at a specific locus on a chromosome may exhibit small nucleotide differences among individuals. Variations in coding or noncoding DNA sequences among a population are called polymorphisms (from the Greek for "many forms").

Pleiotropy: So far, we have treated Mendelian inheritance as though each gene affects only one phenotypic character. Most genes, however, have multiple phenotypic effects, a property called pleiotropy (from the Greek pleion, more).

In humans, for example, pleiotropic alleles are responsible for the multiple symptoms associated with certain hereditary diseases, such as cystic fibrosis and sickle-cell disease, discussed later in this chapter. In the garden pea, the gene that determines flower color also affects the color of the coating on the outer surface of the seed, which can be gray or white. Given the intricate molecular and cellular interactions responsible for an organism's development and physiology, it isn't surprising that a single gene can affect a number of characters.

Notice in Figure 17.6 that there is redundancy in the genetic code, but no ambiguity. For example, although codons GAA and GAG both specify glutamic acid (redundancy), neither of them ever specifies any other amino acid (no ambiguity). The redundancy in the code is not altogether random.

In many cases, codons that are synonyms for a particular amino acid differ only in the third nucleotide base of the triplet. We will consider the significance of this redundancy later in the chapter.

Sister chromatids stay together due to sister chromatid cohesion, mediated by cohesin proteins. In mitosis, this attachment lasts until the end of metaphase, when enzymes cleave the cohesins, freeing the sister chromatids to move to opposite poles of the cell.

In meiosis, sister chromatid cohesion is released in two steps, one at the start of anaphase I and one at anaphase II. In metaphase I, the two homologs of each pair are held together because there is still cohesion between sister chromatid arms in regions beyond points of crossing over, where stretches of sister chromatids now belong to different chromosomes.

Making Multiple Polypeptides in Bacteria and Eukaryotes:

In previous sections, you learned how a single polypeptide is synthesized using the information encoded in an mRNA molecule. When a polypeptide is required in a cell, though, the need is for many copies, not just one.

Genomic Imprinting: Throughout our discussions of Mendelian genetics and the chromosomal basis of inheritance, we have assumed that a given allele will have the same effect whether it was inherited from the mother or the father. This is probably a safe assumption most of the time. For example, when Mendel crossed purple-flowered pea plants with white-flowered pea plants, he observed the same results regardless of whether the purpleflowered parent supplied the eggs or the sperm.

In recent years, however, geneticists have identified a number of traits in mammals that depend on which parent passed along the alleles for those traits. Such variation in phenotype depending on whether an allele is inherited from the male or female parent is called genomic imprinting. (Note that unlike sex-linked genes, most imprinted genes are on autosomes.) Using newer DNA sequence-based methods, about 100 imprinted genes have been identified in humans, and 125 in mice

When the results of Nüsslein-Volhard and Wieschaus were combined with Lewis's earlier work, a coherent picture of Drosophila development emerged.

In recognition of their discoveries, the three researchers were awarded a Nobel Prize in 1995. Next, let's consider a specific example of the genes that Nüsslein-Volhard, Wieschaus, and co-workers found.

Chargaff also noticed a peculiar regularity in the ratios of nucleotide bases. In the DNA of each species he studied, the number of adenines approximately equaled the number of thymines, and the number of guanines approximately equaled the number of cytosines.

In sea urchin DNA, for example, Chargaff's analysis found the four bases in these percentages: A = 32.8% and T = 32.1%; G = 17.7% and C = 17.3%. (The percentages are not exactly the same because of limitations in Chargaff's techniques.)

Origins of Genetic Variation Among Offspring:

In species that reproduce sexually, the behavior of chromosomes during meiosis and fertilization is responsible for most of the variation that arises in each generation. Three mechanisms contribute to the genetic variation arising from sexual reproduction: independent assortment of chromosomes, crossing over, and random fertilization.

Many human traits follow Mendelian patterns of inheritance: Peas are convenient subjects for genetic research, but humans are not. The human generation span is long—about 20 years—and human parents produce many fewer offspring than peas and most other species. Even more important, it wouldn't be ethical to ask pairs of humans to breed so that the phenotypes of their offspring could be analyzed!

In spite of these constraints, the study of human genetics continues, spurred on by our desire to understand our own inheritance and to develop treatments and cures for human genetically based diseases. New molecular biological techniques have led to many breakthrough discoveries, as we will see in Concept 20.4, but basic Mendelian genetics endures as the foundation of human genetics.

The fourth and final part of Mendel's model, the law of segregation, states that the two alleles for a heritable character segregate (separate from each other) during gamete formation and end up in different gametes. Thus, an egg or a sperm gets only one of the two alleles that are present in the somatic cells of the organism making the gamete.

In terms of chromosomes, this segregation corresponds to the distribution of copies of the two members of a pair of homologous chromosomes to different gametes in meiosis (see Figure 13.7).

Several decades later, research supported Garrod's hypothesis that a gene dictates the production of a specific enzyme, later named the one gene-one enzyme hypothesis. Biochemists learned that cells synthesize and degrade most organic molecules via metabolic pathways, in which each chemical reaction in a sequence is catalyzed by a specific enzyme (see Concept 8.1). Such metabolic pathways lead, for instance, to the synthesis of the pigments that give the brown donkey in Figure 17.1 its fur color or fruit flies (Drosophila) their eye color (see Figure 15.3).

In the 1930s, the American biochemist and geneticist George Beadle and his French colleague Boris Ephrussi speculated that in Drosophila, each mutation affecting eye color blocks pigment synthesis at a specific step by preventing production of the enzyme that catalyzes that step. But neither the chemical reactions nor the enzymes that catalyze them were known at the time.

These two findings became known as Chargaff's rules: (1) DNA base composition varies between species, and (2) for each species, the percentages of A and T bases are roughly equal, as are those of G and C bases.

In the Scientific Skills Exercise, you can use Chargaff's rules to predict percentages of nucleotide bases. The basis for these rules remained unexplained until the discovery of the double helix.

Untreated, cystic fibrosis can cause death by the age of 5. Daily doses of antibiotics to stop infection, gentle pounding on the chest to clear mucus from clogged airways, and other therapies can prolong life.

In the United States, more than half of those with cystic fibrosis now survive into their 30s and beyond.

However, the population of daughter cells resulting from meiosis of a large number of diploid cells contains all four types in approximately equal numbers. In the case of n = 3, eight combinations (2 * 2 * 2 = 23 ) of chromosomes are possible for daughter cells. More generally, the number of possible combinations when chromosomes sort independently during meiosis is 2n , where n is the haploid number of the organism.

In the case of humans (n = 23), the number of possible combinations of maternal and paternal chromosomes in the resulting gametes is 223, or about 8.4 million. Each gamete that you produce in your lifetime contains one of roughly 8.4 million possible combinations of chromosomes. This is an underestimate, because it doesn't take into account crossing over, which we'll consider next..

Regulation of both the trp and lac operons involves the negative control of genes because the operons are switched off by the active form of their respective repressor protein. It may be easier to see this for the trp operon, but it is also true for the lac operon.

In the case of the lac operon, allolactose induces enzyme synthesis not by directly activating the lac operon, but by freeing it from the negative effect of the repressor (see Figure 18.4b). Gene regulation is said to be positive only when a regulatory protein interacts directly with the genome to switch transcription on.

In addition to the rarer diseases clearly caused by defects in mitochondrial DNA, mitochondrial mutations inherited from a person's mother may contribute to at least some types of diabetes and heart disease, as well as to other disorders that commonly debilitate the elderly, such as Alzheimer's disease.

In the course of a lifetime, new mutations gradually accumulate in our mitochondrial DNA, and some researchers think that these mutations play a role in the normal aging process.

An mRNA molecule is complementary rather than identical to its DNA template because RNA nucleotides are assembled on the template according to base-pairing rules (see Figure 17.5). The pairs are similar to those that form during DNA replication, except that U (the RNA substitute for T) pairs with A and the mRNA nucleotides contain ribose instead of deoxyribose. Like a new strand of DNA, the RNA molecule is synthesized in an antiparallel direction to the template strand of DNA. (To review what is meant by "antiparallel" and the 5' and 3' ends of a nucleic acid chain, see Figure 16.7.)

In the example in Figure 17.5, the nucleotide triplet ACC along the DNA template strand (written as 3' -ACC-5' ) provides a template for 5'-UGG-3' in the mRNA molecule. The mRNA nucleotide triplets are called codons, and they are customarily written in the 5' -> 3' direction. In our example, UGG is the codon for the amino acid tryptophan (abbreviated Trp, or W). The term codon is also used for the DNA nucleotide triplets along the nontemplate strand. These codons are complementary to the template strand and thus identical in sequence to the mRNA, except that they have a T wherever there is a U in the mRNA. For this reason, the nontemplate DNA strand is often called the coding strand; by convention, the sequence of the coding strand is used when a gene's sequence is reported.

In their experiment, they used a radioactive isotope of sulfur to tag protein in one batch of T2 and a radioactive isotope of phosphorus to tag DNA in a second batch. Because protein, but not DNA, contains sulfur, radioactive sulfur atoms were incorporated only into the protein of the phage. In a similar way, the atoms of radioactive phosphorus labeled only the DNA, not the protein, because nearly all the phage's phosphorus is in its DNA.

In the experiment, separate samples of nonradioactive E. coli cells were infected with the protein-labeled and DNA-labeled batches of T2. The researchers then tested the two samples shortly after the onset of infection to see which type of molecule—protein or DNA—had entered the bacterial cells and would therefore be capable of reprogramming them.

A hallmark of prokaryotic and eukaryotic cells alike—from a bacterium to the cells of a fish—is their intricate and precise regulation of gene expression. In this chapter, we first explore how bacteria regulate expression of their genes in response to different environmental conditions. We then examine the general mechanisms by which eukaryotes regulate gene expression, including the many roles played by RNA molecules.

In the final two sections, we explore the role of gene regulation in both embryonic development, as the ultimate example of proper gene regulation, and cancer, as an illustration of what happens when regulation goes awry. Orchestrating proper gene expression by all cells is crucial to the functions of life.

This abundance of genetic variation provides the raw material on which natural selection works. If the traits conferred by particular combinations of alleles are better suited for a given environment, organisms possessing those genotypes will be expected to thrive and leave more offspring, ensuring the continuation of their genetic complement.

In the next generation, of course, the alleles will be shuffled anew. Ultimately, the interplay between environment and phenotype (and thus genotype) will determine which genetic combinations persist over time.

Some inheritance patterns are exceptions to standard Mendelian inheritance:

In the previous section, you learned about deviations from the usual patterns of chromosomal inheritance due to abnormal events in meiosis and mitosis. We conclude this chapter by describing two normally occurring exceptions to Mendelian genetics, one involving genes located in the nucleus and the other involving genes located outside the nucleus. In both cases, the sex of the parent contributing an allele is a factor in the pattern of inheritance.

Geneticists are working to identify inherited cancer alleles so that predisposition to certain cancers can be detected early in life. About 15% of colorectal cancers, for example, involve inherited mutations. Many of these affect the tumor-suppressor gene called adenomatous polyposis coli, or APC (see Figure 18.26). This gene has multiple functions in the cell, including regulation of cell migration and adhesion. Even in patients with no family history of the disease, the APC gene is mutated in 60% of colorectal cancers.

In these individuals, new mutations must occur in both APC alleles before the gene's function is lost. Since only 15% of colorectal cancers are associated with known inherited mutations, researchers continue to try to identify "markers" that could predict the risk of developing this type of cancer.

Second, cells can adjust the production level of certain enzymes; that is, they can regulate the expression of the genes encoding the enzymes. If, in our example, the environment provides all the tryptophan the cell needs, the cell stops making the enzymes that catalyze the synthesis of tryptophan (Figure 18.2b).

In this case, the control of enzyme production occurs at the level of transcription, the synthesis of messenger RNA from the genes that code for these enzymes.

To a large extent, molecular biology was born in the laboratories of biologists studying viruses that infect bacteria. Experiments with these viruses provided evidence that genes are made of nucleic acids, and they were critical in working out the molecular mechanisms of the fundamental processes of DNA replication, transcription, and translation.

In this chapter, we will explore the biology of viruses, beginning with their structure and then describing how they replicate. Next, we will discuss the role of viruses as diseasecausing agents, or pathogens, and conclude by considering some even simpler infectious agents called prions.

Mendel developed his theory of inheritance several decades before chromosomes were observed under the microscope and the significance of their behavior during mitosis or meiosis was understood.

In this chapter, we'll step into Mendel's garden to re-create his experiments and explain how he arrived at his theory of inheritance. We'll also explore inheritance patterns more complex than those observed by Mendel in garden peas. Finally, we will see how the Mendelian model applies to the inheritance of human variations, including hereditary disorders such as sickle-cell disease.

Fetal Testing: suppose a couple expecting a child learns that they are both carriers of the Tay-Sachs allele. One of the tests that can be done to determine whether the developing fetus has TaySachs disease is amniocentesis, which can be performed starting at the 15th week of pregnancy (Figure 14.19a).

In this procedure, a physician inserts a needle into the uterus and extracts about 10 mL of amniotic fluid, the liquid that bathes the fetus. Some genetic disorders can be detected from the presence of certain molecules in the amniotic fluid itself. Tests for other disorders, including Tay-Sachs disease, are performed on the DNA of cells cultured in the laboratory, descendants of fetal cells sloughed off into the amniotic fluid. A karyotype of these cultured cells can also identify certain chromosomal defects (see Figure 13.3).

Researchers have worked out what happens at the molecular level during muscle cell determination. To do so, they grew embryonic precursor cells in culture and analyzed them using molecular techniques you will learn about in Concepts 20.1 and 20.2. In a series of experiments, they isolated different genes, caused each to be expressed in a separate embryonic precursor cell, and then looked for differentiation into myoblasts and muscle cells.

In this way, they identified several so-called "master regulatory genes" whose protein products commit the cells to becoming skeletal muscle. Thus, in the case of muscle cells, the molecular basis of determination is the expression of one or more of these master regulatory genes.

Consistent with this notion of pieces of DNA shuttling from cell to cell is the observation that a viral genome can have more in common with the genome of its host than with the genomes of viruses that infect other hosts.

Indeed, some viral genes are essentially identical to genes of the host.

An important function of the DNA repair enzymes in our skin cells is to repair genetic damage caused by the ultraviolet rays of sunlight. One type of damage, shown in Figure 16.19, is the covalent linking of thymine bases that are adjacent on a DNA strand. Such thymine dimers cause the DNA to buckle and interfere with DNA replication. The importance of repairing this kind of damage is underscored by a disorder called xeroderma pigmentosum (XP), which in most cases is caused by an inherited defect in a nucleotide excision repair enzyme.

Individuals with XP are hypersensitive to sunlight; mutations in their skin cells caused by ultraviolet light are left uncorrected, often resulting in skin cancer. The effects are extreme: Without sun protection, children who have XP can develop skin cancer by age 10.

The Lysogenic Cycle: Instead of lysing their host cells, many phages coexist with them in a state called lysogeny. In contrast to the lytic cycle, which kills the host cell, the lysogenic cycle allows replication of the phage genome without destroying the host. Phages capable of using both modes of replicating within a bacterium are called temperate phages. A temperate phage called lambda, written with the Greek letter l, has been widely used in biological research. Phage l resembles T4, but its tail has only one short tail fiber.

Infection of an E. coli cell by phage l begins when the phage binds to the surface of the cell and injects its linear DNA genome (Figure 19.6). Within the host, the l DNA molecule forms a circle. What happens next depends on the replicative mode: lytic cycle or lysogenic cycle. During a lytic cycle, the viral genes immediately turn the host cell into a l-producing factory, and the cell soon lyses and releases its virus progeny. During a lysogenic cycle, however, the l DNA molecule is incorporated into a specific site on the E. coli chromosome by viral proteins that break both circular DNA molecules and join them to each other. When integrated into the bacterial chromosome in this way, the viral DNA is known as a prophage.

The researchers dealt with this problem by looking for recessive mutations, which can be propagated in heterozygous flies that act as genetic carriers. Third, cytoplasmic determinants in the egg were known to play a role in axis formation, so the researchers knew they would have to study the mother's genes as well as those of the embryo.

It is the mother's genes that we will discuss further as we focus on how the anteriorposterior body axis is set up in the developing egg.

The island of Asinara lies off the coast of the Italian island of Sardinia. The name Asinara probably originated from the Latin work sinuaria, which means "sinusshaped." A second meaning of Asinara is "donkey-inhabited," which is perhaps even more appropriate because Asinara is home to a wild population of albino donkeys (Figure 17.1). What factors are responsible for the albino phenotype?

Inherited traits are determined by genes, and the trait of albinism is caused by a recessive allele of a pigmentation gene (see Concept 14.4). The information content of genes is in the form of specific sequences of nucleotides along strands of DNA, the genetic material. The albino donkey has a faulty version of a key protein, an enzyme required for pigment synthesis, and this protein is faulty because the gene that codes for it contains incorrect information.

In this chapter, we have seen how sexual reproduction greatly increases the genetic variation present in a population. Although Darwin realized that heritable variation is what makes evolution possible, he could not explain why offspring resemble—but are not identical to—their parents.

Ironically, Gregor Mendel, a contemporary of Darwin, published a theory of inheritance that helps explain genetic variation, but his discoveries had no impact on biologists until 1900, more than 15 years after Darwin (1809-1882) and Mendel (1822-1884) had died. In the next chapter, you'll learn how Mendel discovered the basic rules governing the inheritance of specific traits.

As shown in Figure 16.20, telomeres become shorter during every round of replication. Thus, as expected, telomeric DNA tends to be shorter in dividing somatic cells of older individuals and in cultured cells that have divided many times.

It has been proposed that shortening of telomeres is somehow connected to the aging process of certain tissues and even to aging of the organism as a whole

It appears that this group may not have reproduced sexually for more than 50 million years of their evolutionary history, a model that has been supported by recent analysis of the genetic sequences in its genome. Does this mean that genetic diversity is not advantageous in this species?

It turns out that bdelloid rotifers are an exception to the "rule" that sex alone generates genetic diversity: Bdelloids have mechanisms other than sexual reproduction for generating genetic diversity. For example, they live in environments that can dry up for long periods of time, during which they can enter a state of suspended animation. In this state, their cell membranes may crack in places, allowing entry of DNA from other rotifer species and even from more distantly related species.

Pattern Formation: Setting Up the Body Plan. Cytoplasmic determinants and inductive signals both contribute to the development of a spatial organization in which the tissues and organs of an organism are all in their characteristic places. This process is referred to as pattern formation.

Just as the locations of the front, back, and sides of a new building are determined before construction begins, pattern formation in animals begins in the early embryo, when the major axes of an animal are established. In a bilaterally symmetrical animal, the relative positions of head and tail, right and left sides, and back and front—the three major body axes—are set up before the organs appear. The molecular cues that control pattern formation, collectively called positional information, are provided by cytoplasmic determinants and inductive signals (see Figure 18.17).

The ongoing evolutionary relationship between viruses and the genomes of their host cells is an association that continues to make viruses very useful experimental systems in molecular biology.

Knowledge about viruses also allows many practical applications, since viruses have a tremendous impact on all organisms through their ability to cause disease.

A Borrowed Life: The illustration in Figure 19.1 shows a remarkable event: Human immune cells (purple) infected by human immunodeficiency viruses (HIV) are releasing more HIV viruses. These viruses (red, surrounded by a protein-studded purple membrane from the immune cell) will infect other cells. (The SEM below shows one infected cell.) By injecting its genetic information into a cell, a virus hijacks the cell, recruiting cellular machinery to manufacture many new viruses and promote further infection. Left untreated, HIV causes acquired immunodeficiency syndrome (AIDS) by destroying vital immune system cells. Compared with eukaryotic and even prokaryotic cells, viruses are much smaller and simpler in structure

Lacking the structures and metabolic machinery found in a cell, a virus is an infectious particle consisting of little more than genes packaged in a protein coat.

Large-scale chromosomal changes can also affect an organism's phenotype. Physical and chemical disturbances, as well as errors during meiosis, can damage chromosomes in major ways or alter their number in a cell.

Large-scale chromosomal alterations in humans and other mammals often lead to spontaneous abortion (miscarriage) of a fetus, and individuals born with these types of genetic defects commonly exhibit various developmental disorders. Plants appear to tolerate such genetic defects better than animals do.

Maternal mRNAs are crucial during development of many species. In Drosophila, gradients of specific proteins encoded by maternal mRNAs not only determine the posterior and anterior ends but also establish the dorsal-ventral axis. As the fly embryo grows, it reaches a point when the embryonic program of gene expression takes over, and the maternal mRNAs must be destroyed. (This process involves miRNAs in Drosophila and other species.)

Later, positional information encoded by the embryo's genes, operating on an ever finer scale, establishes a specific number of correctly oriented segments and triggers the formation of each segment's characteristic structures. When the genes operating in this final step are abnormal, the pattern of the adult is abnormal, as you saw in Figure 18.20.

To illustrate the relationship between dominance and phenotype, we can use one of the characters Mendel studied—round versus wrinkled pea seed shape. The dominant allele (round) codes for an enzyme that helps convert an unbranched form of starch to a branched form in the seed. The recessive allele (wrinkled) codes for a defective form of this enzyme, leading to an accumulation of unbranched starch, which causes excess water to enter the seed by osmosis.

Later, when the seed dries, it wrinkles. If a dominant allele is present, no excess water enters the seed and it does not wrinkle when it dries. One dominant allele results in enough of the enzyme to synthesize adequate amounts of branched starch, which means that dominant homozygotes and heterozygotes have the same phenotype: round seeds.

Skin pigmentation in humans is also controlled by many separately inherited genes. Here, we'll simplify the story in order to understand the concept of polygenic inheritance.

Let's consider three genes, with a dark-skin allele for each gene (A, B, or C) contributing one "unit" of darkness (also a simplification) to the phenotype and being incompletely dominant to the other allele (a, b, or c).

The Life Cycle of Drosophila: Fruit flies and other arthropods have a modular construction, an ordered series of segments. These segments make up the body's three major parts: the head, the thorax (the mid-body, from which the wings and legs extend), and the abdomen (Figure 18.19a).

Like other bilaterally symmetrical animals, Drosophila has an anterior-posterior (head-to-tail) axis, a dorsalventral (back-to-belly) axis, and a right-left axis. In Drosophila, cytoplasmic determinants that are localized in the unfertilized egg provide positional information for the placement of anterior-posterior and dorsal-ventral axes even before fertilization. We'll focus here on the molecules involved in establishing the anterior-posterior axis.

Molecular Components of Transcription: Messenger RNA, the carrier of information from DNA to the cell's protein-synthesizing machinery, is transcribed from the template strand of a gene. An enzyme called an RNA polymerase pries the two strands of DNA apart and joins together RNA nucleotides complementary to the DNA template strand, thus elongating the RNA polynucleotide (Figure 17.8).

Like the DNA polymerases that function in DNA replication, RNA polymerases can assemble a polynucleotide only in its 5¿ S 3¿ direction, adding onto its 3¿ end. Unlike DNA polymerases, however, RNA polymerases are able to start a chain from scratch; they don't need to add the first nucleotide onto a pre-existing primer.

Improved DNA-sequencing techniques have transformed the way in which we can explore fundamental biological questions about evolution and how life works (see Make Connections Figure 5.26).

Little more than 15 years after the human genome sequence was announced, researchers had completed the sequencing of thousands of genomes, with tens of thousands in progress. Complete genome sequences have been determined for cells from several cancers, for ancient humans, and for the many bacteria that live in the human intestine. In Chapter 21, you'll learn more about how this rapid acceleration of sequencing technology has revolutionized our study of the evolution of species and the genome itself. Now, let's consider how individual genes are studied.

MITOSIS - Daughter cells of mitosis Prophase: Duplicated chromosome (two sister chromatids) Chromosome duplication 2n = 6 Metaphase: Individual chromosomes line up at the metaphase plate. Anaphase Telophase: Sister chromatids separate during anaphase. Property: Mitosis (occurs in both diploid and haploid cells) DNA replication: Occurs during interphase, before mitosis begins Synapsis of homologous chromosomes: Does not occur. Number of daughter cells and genetic composition: Two, each genetically identical to the parent cell, with the same number of chromosomes Role in animals, fungi, and plants: Enables multicellular animal, fungus, or plant (gametophyte or sporophyte) to arise from a single cell; produces cells for growth, repair, and, in some species, asexual reproduction; produces gametes in the plant gametophyte

MEIOSIS Meiosis I Prophase I: Chiasma (site of crossing over) Pair of duplicated homologs held together by chiasma and sister chromatid cohesion. Chromosome duplication. 2n = 6 Metaphase I: Pairs of homologous chromosomes line up at the metaphase plate. Anaphase I Telophase I: Homologs separate during anaphase I; sister chromatids remain attached at the centromere.Haploid n = 3 MEIOSIS II: Sister chromatids separate during anaphase II. Daughter cells of meiosis II Property: Meiosis (can only occur in diploid cells) DNA Replication: Occurs during interphase before meiosis I but not meiosis II Number of divisions: Two, each including prophase, metaphase, anaphase, and telophase Synapsis of Does not occur homologous chromosomes: Occurs during prophase I along with crossing over between nonsister chromatids; resulting chiasmata hold pairs together due to sister chromatid cohesion. Number of daughter cells and genetic composition: Four, each haploid (n); genetically different from the parent cell and from each other Role in animals, fungi, and plants: Produces gametes (in animals) or spores (in fungi and in plant sporophytes); reduces number of chromosome sets by half and introduces genetic variability among the gametes or spores

The DNA Replication Complex It is traditional—and convenient—to represent DNA polymerase molecules as locomotives moving along a DNA railroad track, but such a model is inaccurate in two important ways. First, the various proteins that participate in DNA replication actually form a single large complex, a "DNA replication machine."

Many protein-protein interactions facilitate the efficiency of this complex. For example, by interacting with other proteins at the fork, primase apparently acts as a molecular brake, slowing progress of the replication fork and coordinating the placement of primers and the rates of replication on the leading and lagging strands. Second, the DNA replication complex may not move along the DNA; rather, the DNA may move through the complex during the replication process. In eukaryotic cells, multiple copies of the complex, perhaps grouped into "factories," may be anchored to the nuclear matrix, a framework of fibers extending through the interior of the nucleus.

Height is a good example of polygenic inheritance: In 2014, a genomic study of over 250,000 people found almost 700 genetic variations associated with over 180 genes that affect height.

Many variations were in or near genes involved in biochemical pathways affecting growth of the skeleton, but others were associated with genes not obviously related to growth.

The Variety of Sexual Life Cycles: Although the alternation of meiosis and fertilization is common to all organisms that reproduce sexually, the timing of these two events in the life cycle varies, depending on the species. These variations can be grouped into three main types of life cycles. In the type that occurs in humans and most other animals, gametes are the only haploid cells (Figure 13.6a).

Meiosis occurs in germ cells during the production of gametes, which undergo no further cell division prior to fertilization. After fertilization, the diploid zygote divides by mitosis, producing a multicellular organism that is diploid.

Mendel used the scientific approach to identify two laws of inheritance:

Mendel discovered the basic principles of heredity by breeding garden peas in carefully planned experiments. As we retrace his work, you will recognize the key elements of the scientific process that were introduced in Chapter 1.

Mendel's Experimental, Quantitative Approach:

Mendel grew up on his parents' small farm in a region of Austria that is now part of the Czech Republic. In this agricultural area, Mendel and the other children received agricultural training in school along with their basic education. As an adolescent, Mendel overcame financial hardship and illness to excel in high school and, later, at the Olmutz Philosophical Institute.

Mendel used very large sample sizes and kept accurate records of his results: 705 of the F2 plants had purple flowers, and 224 had white flowers. These data fit a ratio of approximately three purple to one white (Figure 14.3).

Mendel reasoned that the heritable factor for white flowers did not disappear in the F1 plants but was somehow hidden, or masked, when the purple-flower factor was present. In Mendel's terminology, purple flower color is a dominant trait, and white flower color is a recessive trait. The reappearance of white-flowered plants in the F2 generation was evidence that the heritable factor causing white flowers had not been diluted or destroyed by coexisting with the purple-flower factor in the F1 hybrids. Instead, it had been hidden when in the presence of the purple-flower factor.

Had Mendel stopped his experiments with the F1 generation, the basic patterns of inheritance would have eluded him.

Mendel's quantitative analysis of the F2 plants from thousands of genetic crosses like these allowed him to deduce two fundamental principles of heredity, now called the law of segregation and the law of independent assortment.

Translation is the RNA-directed synthesis of a polypeptide: a closer look We will now examine how genetic information flows from mRNA to protein—the process of translation (Figure 17.15). We'll focus on the basic steps of translation that occur in both bacteria and eukaryotes, while pointing out key differences.

Molecular Components of Translation: In the process of translation, a cell "reads" a genetic message and builds a polypeptide accordingly. The message is a series of codons along an mRNA molecule, and the translator is called a transfer RNA (tRNA). The function of a tRNA is to transfer an amino acid from the cytoplasmic pool of amino acids to a growing polypeptide in a ribosome. A cell keeps its cytoplasm stocked with all 20 amino acids, either by synthesizing them from other compounds or by taking them up from the surrounding solution. The ribosome, a structure made of proteins and RNAs, adds each amino acid brought to it by a tRNA to the growing end of a polypeptide chain (see Figure 17.15).

Eukaryotic DNA Cloning and Expression Systems:

Molecular biologists can avoid eukaryotic-bacterial incompatibility by using eukaryotic cells such as yeasts as hosts for cloning and expressing eukaryotic genes. Yeasts, single-celled fungi, are as easy to grow as bacteria, and they have plasmids, a rarity among eukaryotes.

Inheritance of Organelle Genes: Although our focus in this chapter has been on the chromosomal basis of inheritance, we end with an important amendment: Not all of a eukaryotic cell's genes are located on nuclear chromosomes, or even in the nucleus; some genes are located in organelles in the cytoplasm. Because they are outside the nucleus, these genes are sometimes called extranuclear genes or cytoplasmic genes.

Mitochondria, as well as chloroplasts and other plastids in plants, contain small circular DNA molecules that carry a number of genes. These organelles reproduce themselves and transmit their genes to daughter organelles. Genes on organelle DNA are not distributed to offspring according to the same rules that direct the distribution of nuclear chromosomes during meiosis, so they do not display Mendelian inheritance.

An alternative to the blending model is a "particulate" hypothesis of inheritance: the gene idea. In this model, parents pass on discrete heritable units—genes— that retain their separate identities in offspring. An organism's collection of genes is more like a deck of cards than a pail of paint. Like cards, genes can be shuffled and passed along, generation after generation, in undiluted form.

Modern genetics had its genesis in an abbey garden, where a monk named Gregor Mendel documented a particulate mechanism for inheritance using pea plants (Figure 14.1).

Genomics, Cell Signaling, and Cancer:

Modern medicine that melds genome-wide molecular studies with cell-signaling research is transforming the treatment of many diseases, such as breast cancer. Using microarray analysis (see Figure 20.13) and other techniques, researchers measured the relative levels of mRNA transcripts for every gene in hundreds of breast cancer tumor samples. They identified four major subtypes of breast cancer that differ in their expression of three signal receptors involved in regulating cell growth and division: • Estrogen receptor alpha (ERα) • Progesterone receptor (PR) • HER2, a type of receptor called a receptor tyrosine kinase (see Figure 11.8) (ERα and PR are steroid receptors; see Figure 11.9.) The absence or excess expression of these receptors can cause aberrant cell signaling, leading in some cases to inappropriate cell division, which may contribute to cancer (see Figure 18.24).

The Law of Segregation: If the blending model of inheritance were correct, the F1 hybrids from a cross between purple-flowered and whiteflowered pea plants would have pale purple flowers, a trait intermediate between those of the P generation.

Notice in Figure 14.2 that the experiment produced a very different result: All the F1 offspring had flowers of the same color as the purple-flowered parents

Editing Genes and Genomes: Molecular biologists have long sought techniques for altering, or editing, the genetic material of cells or organisms in a predictable way. In one such technique, called in vitro mutagenesis, specific mutations are introduced into a cloned gene, and the mutated gene is returned to a cell in such a way that it disables ("knocks out") the normal cellular copies of the same gene.

Molecular biologists have long sought techniques for altering, or editing, the genetic material of cells or organisms in a predictable way. In one such technique, called in vitro mutagenesis, specific mutations are introduced into a cloned gene, and the mutated gene is returned to a cell in such a way that it disables ("knocks out") the normal cellular copies of the same gene.

Females with trisomy X (XXX), which occurs once in approximately 1,000 live female births, are healthy and have no unusual physical features other than being slightly taller than average. Triple-X females are at risk for learning disabilities but are fertile.

Monosomy X, which is called Turner syndrome, occurs about once in every 2,500 female births and is the only known viable monosomy in humans. Although these X0 individuals are phenotypically female, they are sterile because their sex organs do not mature. When provided with estrogen replacement therapy, girls with Turner syndrome do develop secondary sex characteristics. Most have normal intelligence.

re usually distributed normally. If either of the aberrant gametes unites with a normal one at fertilization, the zygote will also have an abnormal number of a particular chromosome, a condition known as aneuploidy. Fertilization involving a gamete that has no copy of a particular chromosome will lead to a missing chromosome in the zygote (so that the cell has 2n - 1 chromosomes); the aneuploid zygote is said to be monosomic for that chromosome. If a chromosome is present in triplicate in the zygote (so that the cell has 2n + 1 chromosomes), the aneuploid cell is trisomic for that chromosome. Mitosis will subsequently transmit the anomaly to all embryonic cells.

Monosomy and trisomy are estimated to occur in 10-25% of human conceptions and are the main reason for pregnancy loss. If the organism survives, it usually has a set of traits caused by the abnormal dose of the genes associated with the extra or missing chromosome. Down syndrome is an example of trisomy in humans that will be discussed later. Nondisjunction can also occur during mitosis. If such an error takes place early in embryonic development, then the aneuploid condition is passed along by mitosis to a large number of cells and is likely to have a substantial effect on the organism.

BRCA1 and BRCA2 are considered tumor-suppressor genes because their wild-type alleles protect against breast cancer and their mutant alleles are recessive. (Note that mutations in BRCA1 are commonly found in the genomes of cells from basal-like breast cancers; see Figure 18.27.) The BRCA1 and BRCA2 proteins both appear to function in the cell's DNA damage repair pathway.

More is known about BRCA2, which, along with another protein, helps repair breaks that occur in both strands of DNA; this repair function is crucial for maintaining undamaged DNA. Because DNA breakage can contribute to cancer, it makes sense that the risk of cancer can be lowered by minimizing exposure to DNA-damaging agents, such as the ultraviolet radiation in sunlight and chemicals found in cigarette smoke. Novel genomics-based analyses of specific cancers, such as the approach described in Figure 18.27, are contributing to both early diagnosis and development of treatments that interfere with expression of key genes in tumors. Ultimately, such approaches are expected to lower the death rate from cancer.

The Multistep Model of Cancer Development:

More than one somatic mutation is generally needed to produce all the changes characteristic of a full-fledged cancer cell. This may help explain why the incidence of cancer increases greatly with age. If cancer results from an accumulation of mutations and if mutations occur throughout life, then the longer we live, the more likely we are to develop cancer.

The Search for the Genetic Material: Scientific Inquiry Once T. H. Morgan's group showed that genes exist as parts of chromosomes (described in Concept 15.1), the two chemical components of chromosomes—DNA and protein—emerged as the leading candidates for the genetic material. Until the 1940s, the case for proteins seemed stronger: Biochemists had identified proteins as a class of macromolecules with great heterogeneity and specificity of function, essential requirements for the hereditary material.

Moreover, little was known about nucleic acids, whose physical and chemical properties seemed far too uniform to account for the multitude of specific inherited traits exhibited by every organism. This view gradually changed as the role of DNA in heredity was worked out in studies of bacteria and the viruses that infect them, systems far simpler than fruit flies or humans. Let's trace the search for the genetic material as a case study in scientific inquiry

However, the fact that RNAi can also affect the expression of nonviral cellular genes may reflect a different evolutionary origin for the RNAi pathway.

Moreover, many species, including mammals, apparently produce their own long, double-stranded RNA precursors to small RNAs such as siRNAs. Once produced, these RNAs can interfere with gene expression at stages other than translation, as we'll discuss next.

In the F2 generation, approximately 75% of the seeds were round and 25% were wrinkled—a 3:1 ratio, as in Figure 14.3.

Now let's see how Mendel deduced the law of segregation from his experimental results. In the discussion that follows, we will use modern terms instead of some of the terms used by Mendel. (For example, we'll use "gene" instead of Mendel's "heritable factor.")

The frequency of Down syndrome increases with the age of the mother. While the disorder occurs in just 0.04% (4 out of 10,000 births) of children born to women under age 30, the risk climbs to 0.92% (92 out of 10,000) for mothers at age 40 and is even higher for older mothers. The correlation of Down syndrome with maternal age has not yet been explained.

Most cases result from nondisjunction during meiosis I, and some research points to an age-dependent abnormality in meiosis. Trisomies of some other chromosomes also increase in incidence with maternal age, although infants with other autosomal trisomies rarely survive for long. Medical experts recommend that prenatal screening for trisomies in the embryo be offered to all pregnant women, due to its low risk and useful results. In 2008, a law was passed stipulating that medical practitioners give accurate, up-to-date information about any prenatal or postnatal diagnosis received by parents and that they connect parents with appropriate support services.

How did viruses originate? Viruses have been found that infect every form of life—not just bacteria, animals, and plants, but also archaea, fungi, and algae and other protists. Because they depend on cells for their own propagation, it seems likely that viruses are not the descendants of precellular forms of life but evolved—possibly multiple times—after the first cells appeared.

Most molecular biologists favor the hypothesis that viruses originated from naked bits of cellular nucleic acids that moved from one cell to another, perhaps via injured cell surfaces. The evolution of genes coding for capsid proteins may have allowed viruses to bind cell membranes, thus facilitating the infection of uninjured cells.

Although it is generally agreed that inbreeding causes an increase in autosomal recessive conditions compared to those resulting from matings between unrelated parents, there is debate among geneticists about exactly how much human consanguinity increases the risk of inherited diseases. For one thing, many harmful alleles have such severe effects that a homozygous embryo spontaneously aborts long before birth.

Most societies and cultures have laws or taboos forbidding marriages between close relatives. These rules may have evolved out of empirical observation that in most populations, stillbirths and birth defects are more common when parents are closely related. Social and economic factors have also influenced the development of customs and laws against consanguineous marriages.

Types of Genes Associated with Cancer: The genes that normally regulate cell growth and division during the cell cycle include genes for growth factors, their receptors, and the intracellular molecules of signaling pathways. (To review cell signaling, see Concept 11.2; for regulation of the cell cycle, see Concept 12.3.)

Mutations that alter any of these genes in somatic cells can lead to cancer. The agent of such change can be random spontaneous mutation. However, it is also likely that many cancer-causing mutations result from environmental influences, such as chemical carcinogens, X-rays and other high-energy radiation, and some viruses.

In this chapter, we'll first describe the main techniques for sequencing and manipulating DNA—DNA technology—and for using these DNA tools to analyze gene expression.

Next, we'll explore advances in cloning organisms and producing stem cells, techniques that have both expanded our basic understanding of biology and enhanced our ability to apply that understanding to global problems. In the last section, we'll survey the practical applications of DNA-based biotechnology, the manipulation of organisms or their components to make useful products. Today, the applications of DNA technology affect everything from agriculture to criminal law to medical research. We will end by considering some of the important social and ethical issues that arise as biotechnology becomes more pervasive in our lives.

Transcription is the DNA-directed synthesis of RNA: a closer look

Now that we have considered the linguistic logic and evolutionary significance of the genetic code, we are ready to reexamine transcription, the first stage of gene expression, in greater detail

The Ras protein, encoded by the ras gene (named for rat sarcoma, a connective tissue cancer), is a G protein that relays a signal from a growth factor receptor on the plasma membrane to a cascade of protein kinases (see Figures 11.8 and 11.10). The cellular response at the end of the pathway is the synthesis of a protein that stimulates the cell cycle (Figure 18.24a).

Normally, such a pathway will not operate unless triggered by the appropriate growth factor. But certain mutations in the ras gene can lead to production of a hyperactive Ras protein that triggers the kinase cascade even in the absence of growth factor, resulting in increased cell division (Figure 18.24b). In fact, hyperactive versions or excess amounts of any of the pathway's components can have the same outcome: excessive cell division.

Inheritance patterns are often more complex than predicted by simple Mendelian genetics: In the 20th century, geneticists extended Mendelian principles not only to diverse organisms, but also to patterns of inheritance more complex than those described by Mendel.For the work that led to his two laws of inheritance, Mendel chose pea plant characters that turn out to have a relatively simple genetic basis: Each character is determined by one gene, for which there are only two alleles, one completely dominant and the other completely recessive. (There is one exception: Mendel's pod shape character is actually determined by two genes.)

Not all heritable characters are determined so simply, and the relationship between genotype and phenotype is rarely so straightforward. Mendel himself realized that he could not explain the more complicated patterns he observed in crosses involving other pea characters or other plant species. This does not diminish the utility of Mendelian genetics, however, because the basic principles of segregation and independent assortment apply even to more complex patterns of inheritance. In this section, we will extend Mendelian genetics to hereditary patterns that were not reported by Mendel.

Structure of Viruses: The tiniest viruses are only 20 nm in diameter—smaller than a ribosome. Millions could easily fit on a pinhead. Even the largest known virus, which has a diameter of 1,500 nanometers (1.5 µm), is barely visible under the light microscope. Stanley's discovery that some viruses could be crystallized was exciting and puzzling news.

Not even the simplest of cells can aggregate into regular crystals. But if viruses are not cells, then what are they? Examining the structure of a virus more closely reveals that it is an infectious particle consisting of nucleic acid enclosed in a protein coat and, for some viruses, surrounded by a membranous envelope.

As shown in Figure 13.11, the number of combinations possible for daughter cells formed by meiosis of a diploid cell with two pairs of homologous chromosomes (n = 2) is four: two possible arrangements for the first pair times two possible arrangements for the second pair.

Note that only two of the four combinations of daughter cells shown in the figure would result from meiosis of a single diploid cell, because a single parent cell would have one or the other possible chromosomal arrangement at metaphase I, but not both.

New Combinations of Alleles: Variation for Natural Selection The physical behavior of chromosomes during meiosis contributes to the generation of variation in offspring (see Concept 13.4). Each pair of homologous chromosomes lines up independently of other pairs during metaphase I, and crossing over prior to that, during prophase I, can mix and match parts of maternal and paternal homologs. Mendel's elegant experiments show that the behavior of the abstract entities known as genes—or, more concretely, alleles of genes— also leads to variation in offspring (see Concept 14.1).

Now, putting these different ideas together, you can see that the recombinant chromosomes resulting from crossing over may bring alleles together in new combinations, and the subsequent events of meiosis distribute to gametes the recombinant chromosomes in a multitude of combinations, such as the new variants seen in Figures 15.9 and 15.10. Random fertilization then increases even further the number of variant allele combinations that can be created.

However, the nucleotide-pair substitutions of greatest interest are those that cause a major change in a protein. The alteration of a single amino acid in a crucial area of a protein—such as in the part of the β-globin subunit of hemoglobin shown in Figure 17.26 or in the active site of an enzyme as shown in Figure 8.19— can significantly alter protein activity.

Occasionally, such a mutation leads to an improved protein or one with novel capabilities, but much more often such mutations are neutral or detrimental, leading to a useless or less active protein that impairs cellular function.

The elegant double-helical structure of deoxyribonucleic acid, or DNA, has become an icon of modern biology (Figure 16.1). James Watson and Francis Crick shook the scientific world in April 1953 with their DNA model, which they constructed from sheet metal and wire, shown in the small photo. Gregor Mendel's heritable factors and Thomas Hunt Morgan's genes on chromosomes are, in fact, composed of DNA. Chemically speaking, your genetic endowment is the DNA you inherited from your parents. DNA, the substance of inheritance, is the most celebrated molecule of our time.

Of all nature's molecules, nucleic acids are unique in their ability to direct their own replication from monomers. Indeed, the resemblance of offspring to their parents has its basis in the accurate replication of DNA and its transmission from one generation to the next. Hereditary information in DNA directs the development of your biochemical, anatomical, physiological, and, to some extent, behavioral traits. In this chapter, you will discover how biologists deduced that DNA is the genetic material and how Watson and Crick worked out its structure. You will also learn how a molecule of DNA is copied during DNA replication and how cells repair their DNA. Finally, you will explore how a molecule of DNA is packaged together with proteins in a chromosome.

Rather than modifying histone proteins, a different set of enzymes can methylate the DNA itself on certain bases, usually cytosine. Such DNA methylation occurs in most plants, animals, and fungi. Long stretches of inactive DNA, such as that of inactivated mammalian X chromosomes (see Figure 15.8), are generally more methylated than regions of actively transcribed DNA (although there are exceptions).

On a smaller scale, the DNA of individual genes is usually more heavily methylated in cells in which those genes are not expressed. Removal of the extra methyl groups can turn on some of these genes. Once methylated, genes usually stay that way through successive cell divisions in a given individual. At DNA sites where one strand is already methylated, enzymes methylate the correct daughter strand after each round of DNA replication. Methylation patterns are thus passed on to daughter cells, and cells forming specialized tissues keep a chemical record of what occurred during embryonic development. A methylation pattern maintained in this way also accounts for genomic imprinting in mammals, where methylation permanently regulates expression of either the maternal or paternal allele of particular genes at the start of development (see Figure 15.17).

Today we understand determination in terms of molecular changes. The outcome of determination, observable cell differentiation, is marked by the expression of genes for tissue-specific proteins. These proteins are found only in a specific cell type and give the cell its characteristic structure and function. The first evidence of differentiation is the appearance of mRNAs for these proteins. Eventually, differentiation is observable with a microscope as changes in cellular structure.

On the molecular level, different sets of genes are sequentially expressed in a regulated manner as new cells arise from division of their precursors. A number of the steps in gene expression may be regulated during differentiation, transcription being the most common. In the fully differentiated cell, transcription remains the principal regulatory point for maintaining appropriate gene expression.

Expressing Cloned Eukaryotic Genes:

Once a gene has been cloned in host cells, its protein product can be expressed in large amounts for research or for practical applications, which we'll explore in Concept 20.4. Cloned genes can be expressed in either bacterial or eukaryotic cells; each option has advantages and disadvantages.

Sequential Regulation of Gene Expression During Cellular Differentiation: The earliest changes that set a cell on its path to specialization are subtle ones, showing up only at the molecular level. Before biologists knew much about the molecular changes occurring in embryos, they coined the term determination to refer to the point at which an embryonic cell is irreversibly committed to becoming a particular cell type.

Once it has undergone determination, an embryonic cell can be experimentally placed in another location in the embryo and it will still differentiate into the cell type that is its normal fate. Differentiation, then, is the process by which a cell attains its determined fate. As the tissues and organs of an embryo develop and their cells differentiate, the cells become more noticeably different in structure and function.

The interaction between eukaryotic RNA polymerase II and transcription factors is an example of the importance of protein-protein interactions in controlling eukaryotic transcription.

Once the appropriate transcription factors are firmly attached to the promoter DNA and the polymerase is bound to them in the correct orientation on the DNA, the enzyme unwinds the two DNA strands and begins transcribing the template strand at the start point.

A lethal dominant allele may be passed on, though, if the lethal disease symptoms first appear after reproductive age. In these cases, the individual may already have transmitted the allele to his or her children. For example, a degenerative disease of the nervous system, called Huntington's disease, is caused by a lethal dominant allele that has no obvious phenotypic effect until the individual is about 35 to 45 years old.

Once the deterioration of the nervous system begins, it is irreversible and inevitably fatal. As with other dominant traits, a child born to a parent with the Huntington's disease allele has a 50% chance of inheriting the allele and the disorder (see the Punnett square in Figure 14.18). In the United States, this disease afflicts about one in 10,000 people.

The experimental approaches you have learned about thus far focused on working with molecules, mainly DNA and proteins. In a parallel line of inquiry, biologists have been developing powerful techniques for cloning whole multicellular organisms.

One aim of this work is to obtain special types of cells, called stem cells, that can give rise to all types of tissues. Being able to manipulate stem cells would allow scientists to use the DNA-based methods previously discussed to alter stem cells for the treatment of diseases. Methods involving the cloning of organisms and production of stem cells are the subject of the next section.

Regulation of the tryptophan synthesis pathway is just one example of how bacteria tune their metabolism to changing environments. Many genes of the bacterial genome are switched on or off by changes in the metabolic status of the cell.

One basic mechanism for this control of gene expression in bacteria, described as the operon model, was discovered in 1961 by François Jacob and Jacques Monod at the Pasteur Institute in Paris. Let's see what an operon is and how it works.

Because they control the orientation (polarity) of the egg and consequently that of the fly, these maternal effect genes are also called egg-polarity genes.

One group of these genes sets up the anterior-posterior axis of the embryo, while a second group establishes the dorsal-ventral axis. Like mutations in segmentation genes, mutations in maternal effect genes are generally embryonic lethals.

The Functional and Evolutionary Importance of Introns: Whether or not RNA splicing and the presence of introns have provided selective advantages during evolutionary history is a matter of some debate. In any case, it is informative to consider their possible adaptive benefits. Specific functions have not been identified for most introns, but at least some contain sequences that regulate gene expression, and many affect gene products.

One important consequence of the presence of introns in genes is that a single gene can encode more than one kind of polypeptide. Many genes are known to give rise to two or more different polypeptides, depending on which segments are treated as exons during RNA processing; this is called alternative RNA splicing (see Figure 18.13). Results from the Human Genome Project (discussed in Concept 21.1) suggest that alternative RNA splicing is one reason humans can get along with about the same number of genes as a nematode (roundworm). Because of alternative splicing, the number of different protein products an organism produces can be much greater than its number of genes.

To understand more about how determination occurs in muscle cell differentiation, let's focus on the master regulatory gene called myoD. The myoD gene deserves its designation as a master regulatory gene. Researchers have shown that the MyoD protein it encodes is capable of changing some kinds of fully differentiated nonmuscle cells, such as fat cells and liver cells, into muscle cells. Why doesn't MyoD work on all kinds of cells?

One likely explanation is that activation of musclespecific genes is not solely dependent on MyoD but requires a particular combination of regulatory proteins, some of which are lacking in cells that do not respond to MyoD. The determination and differentiation of other kinds of tissues may play out in a similar fashion. A growing body of experimental evidence supports the idea that master regulatory proteins like MyoD might actually function by opening the chromatin in particular regions. This allows access to transcription machinery for activation of the next set of cell-type-specific genes.

More and more often, next-generation sequencing is complemented (or in some cases replaced) by "third-generation sequencing," with each new technique being faster and less expensive than the previous one. In some new methods, the DNA is neither cut into fragments nor amplified. Instead, a single, very long DNA molecule is sequenced on its own. Several groups have developed techniques that move a single strand of a DNA molecule through a very small pore (a nanopore) in a membrane, identifying the bases one by one by the distinct way each interrupts an electrical current.

One model of this concept is shown in Figure 20.1, in which the pore is a protein channel embedded in a lipid membrane. (Other researchers are using artificial membranes and nanopores.) The idea is that each type of base interrupts the electrical current for a slightly different length of time.

In the cell, eukaryotic DNA is precisely combined with a large amount of protein. Together, this complex of DNA and protein, called chromatin, fits into the nucleus through an elaborate, multilevel system of packing.

Our current view of the successive levels of DNA packing in a chromosome is outlined in Figure 16.22. Study this figure carefully before reading further.

Transcription and translation occur in all organisms. Because most studies have involved bacteria and eukaryotic cells, they are our main focus in this chapter.

Our understanding of transcription and translation in archaea lags behind, but we do know that archaeal cells share some features of gene expression with bacteria and others with eukaryotes.

Researchers have also found a relatively large number of long noncoding RNAs (lncRNAs), ranging from 200 to hundreds of thousands of nucleotides in length, that are expressed at significant levels in specific cell types at particular times.

One such lncRNA is responsible for X chromosome inactivation, which, in most female mammals, prevents expression of genes located on one of the X chromosomes (see Figure 15.8). In this case, lncRNAs—transcripts of the XIST gene located on the chromosome to be inactivated—bind back to and coat that chromosome, and this binding leads to condensation of the entire chromosome into heterochromatin.

Some organisms have more than two complete chromosome sets in all somatic cells. The general term for this chromosomal alteration is polyploidy; the specific terms triploidy (3n) and tetraploidy (4n) indicate three and four chromosomal sets, respectively.

One way a triploid cell may arise is by the fertilization of an abnormal diploid egg produced by nondisjunction of all its chromosomes. Tetraploidy could result from the failure of a 2n zygote to divide after replicating its chromosomes. Subsequent normal mitotic divisions would then produce a 4n embryo.

In the F2 offspring, what color will the flowers be?

One-fourth of the plants have inherited two purple-flower alleles; clearly, these plants will have purple flowers. One-half of the F2 offspring have inherited one purple-flower allele and one white-flower allele; these plants will also have purple flowers, the dominant trait. Finally, onefourth of the F2 plants have inherited two white-flower alleles and will express the recessive trait. Thus, Mendel's model accounts for the 3:1 ratio of traits that he observed in the F2 generation.

A genetic map based on recombination frequencies is called a linkage map. Figure 15.11 shows Sturtevant's linkage map of three genes: the body color (b) and wing size (vg) genes depicted in Figure 15.10 and a third gene, called cinnabar (cn). Cinnabar is one of many Drosophila genes affecting eye color. Cinnabar eyes, a mutant phenotype, are a brighter red than the wild-type color. The recombination frequency between cn and b is 9%; that between cn and vg, 9.5%; and that between b and vg, 17%. In other words, crossovers between cn and b and between cn and vg are about half as frequent as crossovers between b and vg.

Only a map that locates cn about midway between b and vg is consistent with these data, as you can prove to yourself by drawing alternative maps. Sturtevant expressed the distances between genes in map units, defining one map unit as equivalent to a 1% recombination frequency. In practice, the interpretation of recombination data is more complicated than this example suggests. Some genes on a chromosome are so far from each other that a crossover between them is virtually certain. The observed frequency of recombination in crosses involving two such genes can have a maximum value of 50%, a result indistinguishable from that for genes on different chromosomes. In this case, the physical connection between genes on the same chromosome is not reflected in the results of genetic crosses.

Certain sections of a promoter are especially important for binding RNA polymerase in a way that ensures that transcription will begin at the right place. In bacteria, part of the RNA polymerase itself specifically recognizes and binds to the promoter. In eukaryotes, a collection of proteins called transcription factors mediate the binding of RNA polymerase and the initiation of transcription.

Only after transcription factors are attached to the promoter does RNA polymerase II bind to it. The whole complex of transcription factors and RNA polymerase II bound to the promoter is called a transcription initiation complex. Figure 17.9 shows the role of transcription factors and a crucial promoter DNA sequence called the TATA box in forming the initiation complex at a eukaryotic promoter.

Chemical mutagens fall into several categories. Nucleotide analogs are chemicals similar to normal DNA nucleotides but that pair incorrectly during DNA replication.

Other chemical mutagens interfere with correct DNA replication by inserting themselves into the DNA and distorting the double helix. Still other mutagens cause chemical changes in bases that change their pairing properties.

New Mutations and Mutagens: Mutations can arise in a number of ways. Errors during DNA replication or recombination can lead to nucleotide-pair substitutions, insertions, or deletions, as well as to mutations affecting longer stretches of DNA. If an incorrect nucleotide is added to a growing chain during replication, for example, the base on that nucleotide will then be mismatched with the nucleotide base on the other strand. In many cases, the error will be corrected by DNA proofreading and repair systems (see Concept 16.2).

Otherwise, the incorrect base will be used as a template in the next round of replication, resulting in a mutation. Such mutations are called spontaneous mutations. It is difficult to calculate the rate at which such mutations occur. Rough estimates have been made of the rate of mutation during DNA replication for both E. coli and eukaryotes, and the numbers are similar: About one nucleotide in every 1010 is altered, and the change is passed on to the next generation of cells.

Reproductive Cloning of Mammals: In addition to cloning frogs, researchers were able to clone mammals using early embryonic cells as a source of donor nuclei. Until about 20 years ago, though, it was not known whether a nucleus from a fully differentiated cell could be reprogrammed successfully to act as a donor nucleus. In 1997, researchers in Scotland announced the birth of Dolly, a lamb cloned from an adult sheep by nuclear transfer from a differentiated mammary gland cell (Figure 20.17). Using a technique related to that in 20.16, the researchers implanted early embryos into surrogate mothers.

Out of several hundred embryos, one successfully completed normal development, and Dolly was born, a genetic clone of the nucleus donor. At the age of 6, Dolly suffered complications from a lung infection often seen in sheep kept indoors and was euthanized. Another cloned sheep from the same experiment developed an unusual lung disease. This led to speculation that this sheep's cells were in some way not quite as healthy as those of a normal sheep, possibly reflecting incomplete reprogramming of the original transplanted nucleus. Reprogramming involves epigenetic changes that lead to changes in chromatin structure (see Concept 18.2), to be discussed shortly

Using this approach, Nüsslein-Volhard and Wieschaus eventually identified about 1,200 genes essential for pattern formation during embryonic development. Of these, about 120 were essential for normal segmentation.

Over several years, the researchers were able to group these segmentation genes by general function, to find them on the fly's chromosomes, and to isolate many of them for further study in the lab. The result was a detailed molecular understanding of the early steps in pattern formation in Drosophila

Morgan and his students invented a notation for symbolizing alleles in Drosophila that is still widely used for fruit flies. For a given character in flies, the gene takes its symbol from the first mutant (non-wild type) discovered. Thus, the allele for white eyes in Drosophila is symbolized by w. A superscript + identifies the allele for the wild-type trait: w+ for the allele for red eyes, for example.

Over the years, a variety of gene notation systems have been developed for different organisms. For example, human genes are usually written in all capitals, such as HTT for the gene involved in Huntington's disease. (Alleles may use more than one letter, as this one does.)

Mendel's law of segregation.: This diagram shows the genetic makeup of the generations in Figure 14.3. It illustrates Mendel's model for inheritance of the alleles of a single gene. Each plant has two alleles for the gene controlling flower color, one allele inherited from each of the plant's parents. To construct a Punnett square that predicts the F2 generation offspring, we list all the possible gametes from one parent (here, the F1 female) along the left side of the square and all the possible gametes from the other parent (here, the F1 male) along the top. The boxes represent the offspring resulting from all the possible unions of male and female gametes.

P Generation:

Today, researchers also use a DNA polymerase from the archaean species Pyrococcus furiosus. This enzyme, called Pfu polymerase, is more accurate and stable but more expensive than Taq polymerase.

PCR is speedy and very specific. Only a minuscule amount of DNA need be present in the starting material, and this DNA can be partially degraded, as long as there are a few copies of the complete target sequence. The key to the high specificity is the pair of primers used for each PCR amplification. The primer sequences are chosen so they hybridize only to sequences at opposite ends of the target segment, one on the 3¿ end of each strand. (For high specificity, the primers must be at least 15 or so nucleotides long.) With each successive cycle, the number of target segment molecules of the correct length doubles, so the number of molecules equals 2n , where n is the number of cycles. After 30 or so cycles, about a billion copies of the target sequence are present!

Ultrasound and isolation of fetal cells or DNA from maternal blood pose no known risk to either mother or fetus, while the other procedures can cause complications in a small percentage of cases. Amniocentesis or CVS for diagnostic testing is generally offered to women over age 35, due to their increased risk of bearing a child with Down syndrome, and may also be offered to younger women if there are known concerns. If the fetal tests reveal a serious disorder like Tay-Sachs, the parents face the difficult choice of either terminating the pregnancy or preparing to care for a child with a genetic disorder, one that might even be fatal.

Parental and fetal screening for Tay-Sachs alleles done since 1980 has reduced the number of children born with this incurable disease by 90%. In 2008, the Chinese government initiated a program of fetal testing to detect a harmful genetic blood disorder called β-thalassemia. This effort resulted in a reduction in the rate of this disorder from over 21 births per 1000 in 2008 to just under 13 in 2011.

Inheritance of Genes

Parents endow their offspring with coded information in the form of hereditary units called genes. The genes we inherit from our mothers and fathers are our genetic link to our parents, and they account for family resemblances such as shared eye color or freckles. Our genes program specific traits that emerge as we develop from fertilized eggs into adults.

The debate about the origin of viruses was reinvigorated about 15 years ago by reports of one of the largest viruses yet discovered: Mimivirus is a double-stranded DNA (dsDNA) virus with an icosahedral capsid that is 400 nm in diameter, the size of a small bacterium. Its genome contains 1.2 million bases (Mb)—about 100 times as many as the influenza virus genome—and an estimated 1,000 genes.

Perhaps the most surprising aspect of mimivirus, however, was that its genome included genes previously found only in cellular genomes.

Evidence That Viral DNA Can Program Cells: Additional evidence that DNA was the genetic material came from studies of viruses that infect bacteria (Figure 16.3). These viruses are called bacteriophages (meaning "bacteria-eaters"), or phages for short. Viruses are much simpler than cells. A virus is little more than DNA (or sometimes RNA) enclosed by a protective coat, which is often simply protein. To produce more viruses, a virus must infect a cell and take over the cell's metabolic machinery.

Phages have been widely used as tools by researchers in molecular genetics. In 1952, Alfred Hershey and Martha Chase performed xperiments showing that DNA is the genetic material of a phage known as T2. This is one of many phages that infect Escherichia coli (E. coli), a bacterium that normally lives in the intestines of mammals and is a model organism for molecular biologists. At that time, biologists already knew that T2, like many other phages, was composed almost entirely of DNA and protein. They also knew that the T2 phage could quickly turn an E. coli cell into a T2-producing factory that released many copies of new phages when the cell ruptured.

Cloning Plants: Single-Cell Cultures: The successful cloning of whole plants from single differentiated cells was accomplished during the 1950s by F. C. Steward and his students at Cornell University, who worked with carrot plants. They found that differentiated cells taken from the root (the carrot) and incubated in culture medium could grow into normal adult plants, each genetically identical to the parent plant. These results showed that differentiation does not necessarily involve irreversible changes in the DNA. In plants, mature cells can "dedifferentiate" and then give rise to all the specialized cell types of the organism. Any cell with this potential is said to be totipotent

Plant cloning is used extensively in agriculture. For plants such as orchids, cloning is the only commercially practical means of reproducing plants. In other cases, cloning has been used to reproduce a plant with valuable characteristics, such as resistance to plant pathogens. In fact, you yourself may be a plant cloner: If you have ever grown a new plant from a cutting, you have practiced cloning!

What determines whether a ribosome is free in the cytosol or bound to rough ER?

Polypeptide synthesis always begins in the cytosol as a free ribosome starts to translate an mRNA molecule. There, the process continues to completion—unless the growing polypeptide itself cues the ribosome to attach to the ER.

Synthesizing a New DNA Strand: The unwound sections of parental DNA strands are now available to serve as templates for the synthesis of new complementary DNA strands. However, the enzymes that synthesize DNA cannot initiate the synthesis of a polynucleotide; they can only add DNA nucleotides to the end of an already existing chain that is base-paired with the template strand. The initial nucleotide chain that is produced during DNA synthesis is actually a short stretch of RNA, not DNA. This RNA chain is called a primer and is synthesized by the enzyme primase (see Figure 16.13).

Primase starts a complementary RNA chain with a single RNA nucleotide and adds RNA nucleotides one at a time, using the parental DNA strand as a template. The completed primer, generally 5-10 nucleotides long, is thus base-paired to the template strand. The new DNA strand will start from the 3′ end of the RNA primer. Enzymes called DNA polymerases catalyze the synthesis of new DNA by adding nucleotides to the 3′ end of a preexisting chain. In E. coli, there are several DNA polymerases, but two appear to play the major roles in DNA replication: DNA polymerase III and DNA polymerase I. The situation in eukaryotes is more complicated, with at least 11 different DNA polymerases discovered so far, although the general principles are the same.

To give two examples, the calculations for finding the probabilities of two of the possible F2 genotypes (YYRR and YyRR) are shown below: Probability of YYRR = 1/4 (probability of YY)× 1/4 (RR) = 1/16.

Probability of YyRR = 1/2 (Yy) X 1/4 (RR) =1/8. The YYRR genotype corresponds to the upper left box in the larger Punnett square in Figure 14.8 (one box = 1 ⁄ 16). Looking closely at the larger Punnett square in Figure 14.8, you will see that 2 of the 16 boxes (1 ⁄ 8) correspond to the YyRR genotype.

Figure 13.12 The results of crossing over during meiosis.: Pair of homologs Chiasma, site of crossing over Centromere Daughter cells Recombinant chromosomes

Prophase I of meiosis: In prophase I, synapsis and crossing over occur; then homologs move apart slightly. Chiasmata and cohesion between sister chromatids hold homologs together; they move to the metaphase I plate. Breakdown of proteins holding sister chromatid arms together allows homologs with recombinant chromatids to separate.

Hemophilia is an X-linked recessive disorder defined by the absence of one or more of the proteins required for blood clotting. When a person with hemophilia is injured, bleeding is prolonged because a firm clot is slow to form. Small cuts in the skin are usually not a problem, but bleeding in the muscles or joints can be painful and can lead to serious damage. In the 1800s, hemophilia was widespread among the royal families of Europe.

Queen Victoria of England is known to have passed the allele to several of her descendants. Subsequent intermarriage with royal family members of other nations, such as Spain and Russia, further spread this X-linked trait, and its incidence is well documented in royal pedigrees. A few years ago, new genomic techniques allowed sequencing of DNA from tiny amounts isolated from the buried remains of royal family members. The genetic basis of the mutation, and how it resulted in a nonfunctional blood-clotting factor, is now understood. Today, people with hemophilia are treated as needed with intravenous injections of the protein that is missing.

Organization of a Typical Eukaryotic Gene and Its Transcript: A eukaryotic gene and the DNA elements (segments) that control it are typically organized as shown in Figure 18.8, which extends what you learned about eukaryotic genes in Chapter 17. Recall that a cluster of proteins called a transcription initiation complex assembles on the promoter sequence at the "upstream" end of the gene (see Figure 17.9). One of these proteins, RNA polymerase II, then proceeds to transcribe the gene, synthesizing a primary RNA transcript (premRNA).

RNA processing includes enzymatic addition of a 5′ cap and a poly-A tail, as well as splicing out of introns, to yield a mature mRNA. Associated with most eukaryotic genes are multiple control elements, segments of noncoding DNA that serve as binding sites for the proteins called transcription factors, which bind to the control elements and regulate transcription. Control elements on the DNA and the transcription factors that bind to them are critical to the precise regulation of gene expression seen in different cell types.

Increasingly, with the advent of rapid, inexpensive DNA sequencing methods, microarray usage is decreasing: Researchers can now afford to simply sequence the cDNA samples from different tissues or different embryonic stages in order to discover which genes are expressed. This straightforward method is called RNA sequencing, or RNA-seq (pronounced "RNA-seek"), even though it is the cDNA that is actually sequenced. In RNA-seq, the mRNA (or other RNA) samples are isolated, cut into shorter, similar-sized fragments, and converted into cDNAs (Figure 20.13). These short cDNA stretches are sequenced, and a computer program reassembles them, either mapping them onto the genome of the species in question (when available) or simply ordering them from scratch based on overlapping sequences of multiple RNAs

RNA-seq has several advantages over microarrays. First, the procedure is not based on hybridization with a labeled probe, so it doesn't depend on having genomic sequences in hand (although they are usually available). Second, it can measure levels of expression over a very wide range, unlike microarrays, which cannot accurately measure either very low or very high levels. Third, a careful analysis provides a wealth of information about expression of a particular gene, such as relative levels of alternatively spliced mRNAs. As the price of DNA sequencing plummets, RNA-seq is becoming more widely used for many applications. In most cases, however, expression of individual genes still needs to be confirmed by RT-PCR.

Other mRNA detection techniques may be preferable for comparing the amounts of a specific mRNA in several samples at the same time—for example, in different cell types or in embryos at different stages of development. One method that is widely used is called the reverse transcriptase polymerase chain reaction, or RT-PCR.

RT-PCR begins by turning sample sets of mRNAs into double-stranded DNAs with the corresponding sequences. First, the enzyme reverse transcriptase (from a retrovirus; see Figure 19.9) is used to synthesize a complementary DNA copy of each mRNA in the sample, called a reverse transcript (Figure 20.10). Recall that the 3¿ end of an mRNA has a stretch of adenine (A) nucleotides called a poly-A tail. This allows a short complementary strand of thymine deoxyribonucleotides (poly-dT) to be added and used as a primer for synthesis of this DNA strand.

Effects on mRNAs by MicroRNAs and Small Interfering RNAs:

Regulation by both small and large ncRNAs occurs at several points in the pathway of gene expression, including mRNA translation and chromatin modification. We'll examine two types of small ncRNAs, the importance of which was acknowledged when their discovery was the focus of the 2006 Nobel Prize in Physiology or Medicine, which was awarded for work completed only eight years earlier.

The Behavior of Recessive Alleles: How can we account for the behavior of alleles that cause recessively inherited disorders?

Recall that genes code for proteins of specific function. An allele that causes a genetic disorder (let's call it allele a) codes for either a malfunctioning protein or no protein at all. In the case of disorders classified as recessive, heterozygotes (Aa) typically have the normal phenotype because one copy of the normal allele (A) produces a sufficient amount of the specific protein. Thus, a recessively inherited disorder shows up only in the homozygous individuals (aa) who inherit a recessive allele from each parent. Although phenotypically normal with regard to the disorder, heterozygotes may transmit the recessive allele to their offspring and thus are called carriers. Figure 14.16 illustrates these ideas using albinism as an example.

The Stages of Meiosis: The overview of meiosis in Figure 13.7 shows, for a single pair of homologous chromosomes in a diploid cell, that both members of the pair are duplicated and the copies sorted into four haploid daughter cells

Recall that sister chromatids are two copies of one chromosome, closely associated all along their lengths; this association is called sister chromatid cohesion. Together, the sister chromatids make up one duplicated chromosome (see Figure 13.4).

Protein Processing and Degradation: The final opportunities for controlling gene expression occur after translation. Often, eukaryotic polypeptides must be processed to yield functional protein molecules. For instance, cleavage of the initial insulin polypeptide (pro-insulin) forms the active hormone. In addition, many proteins undergo chemical modifications that make them functional.

Regulatory proteins are commonly activated or inactivated by the reversible addition of phosphate groups (see Figure 11.10), and proteins destined for the surface of animal cells acquire sugars (see Figure 6.12). Cell-surface proteins and many others must also be transported to target destinations in the cell in order to function (see Figure 17.22). Regulation might occur at any of the steps involved in modifying or transporting a protein

RNA Processing: RNA processing in the nucleus and the export of mature RNA to the cytoplasm provide opportunities for regulating gene expression not available in prokaryotes. One example of regulation at the RNA-processing level is alternative RNA splicing, in which different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns.

Regulatory proteins specific to a cell type control intron-exon choices by binding to regulatory sequences within the primary transcript.

Getting Started: The replication of chromosomal DNA begins at particular sites called origins of replication, short stretches of DNA that have a specific sequence of nucleotides. The E. coli chromosome, like many other bacterial chromosomes, is circular and has a single origin. Proteins that initiate DNA replication recognize this sequence and attach to the DNA, separating the two strands and opening up a replication "bubble" (Figure 16.12a).

Replication of DNA then proceeds in both directions until the entire molecule is copied. In contrast to a bacterial chromosome, a eukaryotic chromosome may have hundreds or even a few thousand replication origins. Multiple replication bubbles form and eventually fuse, thus speeding up the copying of the very long DNA molecules (Figure 16.12b). As in bacteria, eukaryotic DNA replication proceeds in both directions from each origin.

In the context of gene regulation, the enzymes of the lactose pathway are referred to as inducible enzymes because their synthesis is induced by a chemical signal (allolactose, in this case). Analogously, the enzymes for tryptophan synthesis are said to be repressible.

Repressible enzymes generally function in anabolic pathways, which synthesize essential end products from raw materials (precursors). By suspending production of an end product when it is already present in sufficient quantity, the cell can allocate its organic precursors and energy for other uses. In contrast, inducible enzymes usually function in catabolic pathways, which break down a nutrient to simpler molecules. By producing the appropriate enzymes only when the nutrient is available, the cell avoids wasting energy and precursors making proteins that are not needed.

Epigenetic Inheritance: The chromatin modifications that we just discussed do not change the DNA sequence, yet they still may be passed along to future generations of cells. Inheritance of traits transmitted by mechanisms not involving the nucleotide sequence itself is called epigenetic inheritance. Whereas mutations in the DNA are permanent changes, modifications to the chromatin can be reversed. For example, DNA methylation patterns are largely erased and reestablished during gamete formation.

Researchers are amassing more and more evidence for the importance of epigenetic information in the regulation of gene expression. Epigenetic variations might help explain why one identical twin acquires a genetically based disease, such as schizophrenia, but the other does not, despite their identical genomes. Alterations in normal patterns of DNA methylation are seen in some cancers, where they are associated with inappropriate gene expression. Evidently, enzymes that modify chromatin structure are integral parts of the eukaryotic cell's machinery for regulating transcription

DNA Sequencing:

Researchers can exploit the principle of complementary base pairing to determine the complete nucleotide sequence of a DNA molecule, a process called DNA sequencing. The DNA is first cut into fragments, and then each fragment is sequenced. The first automated procedure used a technique called dideoxyribonucleotide (or dideoxy) chain termination sequencing. In this technique, one strand of a DNA fragment is used as a template for synthesis of a nested set of complementary fragments; these are further analyzed to yield the sequence. Biochemist Frederick Sanger received the Nobel Prize in 1980 for developing this method. Dideoxy sequencing is still used for routine small-scale sequencing jobs

In eukaryotes, the rate of gene expression can be strongly increased or decreased by the binding of specific transcription factors, either activators or repressors, to the control elements of enhancers. Hundreds of transcription activators have been discovered in eukaryotes; the structure of one example is shown in Figure 18.9.

Researchers have identified two types of structural domains that are commonly found in a large number of activator proteins: a DNA-binding domain—a part of the protein's three-dimensional structure that binds to DNA— and one or more activation domains. Activation domains bind other regulatory proteins or components of the transcription machinery, facilitating a series of protein-protein interactions that result in enhanced transcription of a given gene.

Faulty Gene Regulation in Cloned Animals Due to Epigenetic Differences: In most nuclear transplantation studies thus far, only a small percentage of cloned embryos develop normally to birth. And like Dolly, many cloned animals exhibit defects. Cloned mice, for instance, are prone to obesity, pneumonia, liver failure, and premature death. Scientists assert that even cloned animals that appear normal are likely to have subtle defects.

Researchers have uncovered some reasons for the low efficiency of cloning and the high incidence of abnormalities. In the nuclei of fully differentiated cells, a small subset of genes is turned on and expression of the rest of the genes is repressed. This regulation often is the result of epigenetic changes in chromatin, such as acetylation of histones or methylation of DNA (see Figure 18.7). During the nuclear transfer procedure, many of these changes must be reversed in the laterstage nucleus from a donor animal for genes to be expressed or repressed appropriately in early stages of development. Researchers have found that the DNA in cells from cloned embryos, like that of differentiated cells, often has more methyl groups than does the DNA in equivalent cells from normal embryos of the same species. This finding suggests that the reprogramming of donor nuclei requires more accurate and complete chromatin restructuring than occurs during cloning procedures. Because DNA methylation helps regulate gene expression, misplaced or extra methyl groups in the DNA of donor nuclei may interfere with the pattern of gene expression necessary for normal embryonic development. In fact, the success of a cloning attempt may depend in large part on whether or not the chromatin in the donor nucleus can be artificially modified to resemble that of a newly fertilized egg.

The p53 gene has been called the "guardian angel of the genome." Once the gene is activated—for example, by DNA damage—the p53 protein functions as an activator for several other genes. Often it activates a gene called p21, whose product halts the cell cycle by binding to cyclindependent kinases, allowing time for the cell to repair the DNA.

Researchers recently showed that p53 also activates expression of a group of miRNAs, which in turn inhibit the cell cycle. In addition, the p53 protein can turn on genes directly involved in DNA repair. Finally, when DNA damage is irreparable, p53 activates "suicide" genes, whose protein products bring about programmed cell death (apoptosis; see Figure 11.20). Thus, p53 acts in several ways to prevent a cell from passing on mutations due to DNA damage. If mutations do accumulate and the cell survives through many divisions—as is more likely if the p53 tumor-suppressor gene is defective or missing—cancer may ensue. The many functions of p53 suggest a complex picture of regulation in normal cells, one that we do not yet fully understand.

Inquiry :When F1 hybrid pea plants self- or crosspollinate, which traits appear in the F2 generation? Experiment : Mendel crossed true-breeding purple-flowered plants and white-flowered plants (crosses are symbolized by *). The resulting F1 hybrids were allowed to self-pollinate or were cross-pollinated with other F1 hybrids. The F2 generation plants were then observed for flower color. P Generation (True breeding parents): Purple & white F 1 Generation is the hybrid: all plants have purple flowers self or cross pollination F2 Generation: 705 purple-flowered plants. 224 white-flowered plants

Results: Both purple-flowered and white-flowered plants appeared in the F2 generation, in a ratio of approximately 3:1. Conclusion: The "heritable factor" for the recessive trait (white flowers) had not been destroyed, deleted, or "blended" in the F1 generation but was merely masked by the presence of the factor for purple flowers, which is the dominant trait.

The widely accepted model is that rRNAs, rather than ribosomal proteins, are primarily responsible for both the structure and the function of the ribosome. The proteins, which are largely on the exterior, support the shape changes of the rRNA molecules as they carry out catalysis during translation.

Ribosomal RNA is the main constituent of the A and P sites and of the interface between the two subunits; it also acts as the catalyst of peptide bond formation. Thus, a ribosome could actually be considered one colossal ribozyme!

The Structure and Function of Ribosomes:

Ribosomes facilitate the specific coupling of tRNA anticodons with mRNA codons during protein synthesis. A ribosome consists of a large subunit and a small subunit, each made up of proteins and one or more ribosomal RNAs (rRNAs).

A virus consists of a nucleic acid surrounded by a protein coat:

Scientists were able to detect viruses indirectly long before they were actually able to see them. The story of how viruses were discovered begins near the end of the 19th century.

The bicoid research was groundbreaking for several reasons. First, it led to the identification of a specific protein required for some of the earliest steps in pattern formation. It thus helped us understand how different regions of the egg can give rise to cells that go down different developmental pathways.

Second, it increased our understanding of the mother's critical role in the initial phases of embryonic development. Finally, the principle that a gradient of morphogens can determine polarity and position has proved to be a key developmental concept for a number of species, just as early embryologists had hypothesized

Telomeres have two protective functions. First, specific proteins associated with telomeric DNA prevent the staggered ends of the daughter molecule from activating the cell's systems for monitoring DNA damage. (Staggered ends of a DNA molecule, which often result from double-strand breaks, can trigger signal transduction pathways leading to cell cycle arrest or cell death.)

Second, telomeric DNA acts as a kind of buffer zone that provides some protection against the organism's genes shortening, somewhat like how the plastic-wrapped ends of a shoelace slow down its unraveling. Telomeres do not prevent the erosion of genes near the ends of chromosomes; they merely postpone it.

Meosis I

Separates homologous chromosomes

Sickle-Cell Disease: A Genetic Disorder with Evolutionary Implications: The most common inherited disorder among people of African descent is sickle-cell disease, which affects one out of 400 African-Americans. Sickle-cell disease is caused by the substitution of a single amino acid in the hemoglobin protein of red blood cells; in homozygous individuals, all hemoglobin is of the sickle-cell (abnormal) variety. When the oxygen content of an affected individual's blood is low (at high altitudes or under physical stress, for instance), the sickle-cell hemoglobin proteins aggregate into long fibers that deform the red cells into a sickle shape (see Figure 5.19).

Sickled cells may clump and clog small blood vessels, often leading to other symptoms throughout the body, including physical weakness, pain, organ damage, and even stroke and paralysis. Regular blood transfusions can ward off brain damage in children with sickle-cell disease, and new drugs can help prevent or treat other problems. There is currently no widely available cure, but the disease is the target of ongoing gene therapy research.

Over the past 10 years, biologists have developed a powerful new technique for gene editing in living cells and organisms, called the CRISPR-Cas9 system, that is taking the field of genetic engineering by storm. Cas9 is a bacterial protein that helps defend bacteria against bacteriophage infections in a system worked out by Jennifer Doudna and Emmanuelle Charpentier. In bacterial cells, Cas9 acts together with a "guide RNA" made from the CRISPR region of the bacterial system (see Figure 19.7).

Similar to the restriction enzymes described earlier, Cas9 is a nuclease that cuts double-stranded DNA molecules. However, while a given restriction enzyme recognizes only one particular DNA sequence, the Cas9 protein will cut any sequence to which it is directed. Cas9 takes its marching orders from a guide RNA molecule that it binds and uses as a homing device, cutting both strands of any DNA sequence that is exactly complementary to the guide RNA.

In addition to influencing transcription directly, some activators and repressors act indirectly by affecting chromatin structure. Studies using yeast and mammalian cells show that some activators recruit proteins that acetylate histones near the promoters of specific genes, thus promoting transcription (see Figure 18.7).

Similarly, some repressors recruit proteins that remove acetyl groups from histones, leading to reduced transcription, a phenomenon referred to as silencing. Indeed, recruitment of chromatin-modifying proteins seems to be the most common mechanism of repression in eukaryotic cells

The term phenotype can refer not only to specific characters, such as flower color and blood group, but also to an organism in its entirety—all aspects of its physical appearance, internal anatomy, physiology, and behavior.

Similarly, the term genotype can refer to an organism's entire genetic makeup, not just its alleles for a single genetic locus. In most cases, a gene's impact on phenotype is affected by other genes and by the environment. In this integrated view of heredity and variation, an organism's phenotype reflects its overall genotype and unique environmental history.

Differentiated cells are specialists at making tissue-specific proteins. For example, as a result of transcriptional regulation, liver cells specialize in making albumin, and lens cells specialize in making crystallin (see Figure 18.11). Skeletal muscle cells in vertebrates are another instructive example. Each of these cells is a long fiber containing many nuclei within a single plasma membrane.

Skeletal muscle cells have high concentrations of muscle-specific versions of the contractile proteins myosin and actin, as well as membrane receptor proteins that detect signals from nerve cells.

The biochemical, physiological, and anatomical features associated with "males" and "females" are turning out to be more complicated than previously thought, with many genes involved in their development. Because of the complexity of this process, many variations exist.

Some individuals are born with intermediate sexual ("intersex") characteristics, or even with anatomical features that do not match an individual's sense of their own gender ("transgender" individuals). Sex determination is an active area of research that should yield a more sophisticated understanding in years to come.

Specific transcription factors that function as repressors can inhibit gene expression in several different ways.

Some repressors bind directly to control element DNA (in enhancers or elsewhere), blocking activator binding. Other repressors interfere with the activator itself so it can't bind the DNA.

Each particular virus can infect cells of only a limited number of host species, called the host range of the virus. This host specificity results from the evolution of recognition systems by the virus. Viruses usually identify host cells by a "lock-and-key" fit between viral surface proteins and specific receptor molecules on the outside of cells. According to one model, such receptor molecules originally carried out functions that benefited the host cell but were co-opted later by viruses as portals of entry.

Some viruses have broad host ranges. For example, West Nile virus and equine encephalitis virus are distinctly different viruses that can each infect mosquitoes, birds, horses, and humans. Other viruses have host ranges so narrow that they infect only a single species. Measles virus, for instance, can infect only humans. Furthermore, viral infection of multicellular eukaryotes is usually limited to particular tissues. Human cold viruses infect only the cells lining the upper respiratory tract, and the HIV seen in Figure 19.1 binds to receptors present only on certain types of immune cells.

A third type of life cycle occurs in most fungi and some protists, including some algae (Figure 13.6c). After gametes fuse and form a diploid zygote, meiosis occurs without a multicellular diploid offspring developing. Meiosis produces not gametes but haploid cells that then divide by mitosis and give rise to either unicellular descendants or a haploid multicellular adult organism

Subsequently, the haploid organism carries out further mitoses, producing the cells that develop into gametes. The only diploid stage found in these species is the single-celled zygote.

Substitutions: A nucleotide-pair substitution is the replacement of one nucleotide and its partner with another pair of nucleotides (Figure 17.27a). Some substitutions have no effect on the encoded protein, owing to the redundancy of the genetic code. For example, if 3'-CCG-5' on the template strand mutated to 3'-CCA-5', the mRNA codon that used to be GGC would become GGU, but a glycine would still be inserted at the proper location in the protein (see Figure 17.6). In other words, a change in a nucleotide pair may transform one codon into another that is translated into the same amino acid.

Such a change is an example of a silent mutation, which has no observable effect on the phenotype. (Silent mutations can occur outside genes as well.) Interestingly, there is evidence that some silent mutations may indirectly affect where or at what level the gene gets expressed, even though the actual protein is the same.

Substitutions that change one amino acid to another one are called missense mutations.

Such a mutation may have little effect on the protein: The new amino acid may have properties similar to those of the amino acid it replaces, or it may be in a region of the protein where the exact sequence of amino acids is not essential to the protein's function.

Insertions and Deletions: Insertions and deletions are additions or losses of nucleotide pairs in a gene (Figure 17.27b). These mutations have a disastrous effect on the resulting protein more often than substitutions do. Insertion or deletion of nucleotides may alter the reading frame of the genetic message, the triplet grouping of nucleotides on the mRNA that is read during translation.

Such a mutation, called a frameshift mutation, occurs whenever the number of nucleotides inserted or deleted is not a multiple of three. All nucleotides downstream of the deletion or insertion will be improperly grouped into codons; the result will be extensive missense mutations, usually ending sooner or later in a nonsense mutation that leads to premature termination. Unless the frameshift is very near the end of the gene, the protein is almost certain to be nonfunctional. Insertions and deletions also occur outside of coding regions; these are not called frameshift mutations, but can have effects on the phenotype— for instance, they can affect how a gene is expressed.

Chromosomal translocations can also occur during mitosis; some have been implicated in certain cancers, including chronic myelogenous leukemia (CML). This disease occurs when a reciprocal translocation happens during mitosis of cells that are precursors of white blood cells. The exchange of a large portion of chromosome 22 with a small fragment from a tip of chromosome 9 produces a much shortened, easily recognized chromosome 22, called the Philadelphia chromosome (Figure 15.16).

Such an exchange causes cancer by creating a new "fused" gene that leads to uncontrolled cell cycle progression. (The mechanism of gene activation will be discussed in Chapter 18.)

After the viral nucleic acid molecules and capsomeres are produced, they spontaneously self-assemble into new viruses. In fact, researchers can separate the RNA and capsomeres of TMV and then reassemble complete viruses simply by mixing the components together under the right conditions. The simplest type of viral replicative cycle ends with the exit of hundreds or thousands of viruses from the infected host cell, a process that often damages or destroys the cell.

Such cellular damage and death, as well as the body's responses to this destruction, cause many of the symptoms associated with viral infections. The viral progeny that exit a cell have the potential to infect additional cells, spreading the viral infection. There are many variations on the simplified viral replicative cycle we have just described. We will now take a look at some of these variations in bacterial viruses (phages) and animal viruses; later in the chapter, we will consider plant viruses.

Cloning Animals: Nuclear Transplantation Differentiated cells from animals generally do not divide in culture, much less develop into the multiple cell types of a new organism. Therefore, early researchers had to use a different approach to answer the question of whether differentiated animal cells are totipotent. Their approach was to remove the nucleus of an egg (creating an enucleated egg) and replace it with the nucleus of a differentiated cell, a procedure called nuclear transplantation, now more commonly called somatic cell nuclear transfer. If the nucleus from the differentiated donor cell retains its full genetic capability, then it should be able to direct development of the recipient cell into all the tissues and organs of an organism.

Such experiments were conducted on one species of frog (Rana pipiens) by Robert Briggs and Thomas King in the 1950s and on another frog species (Xenopus laevis) by John Gurdon in the 1970s (Figure 20.16). These researchers transplanted a nucleus from an embryonic or tadpole cell into an enucleated egg of the same species. In Gurdon's experiments, the transplanted nucleus was often able to support normal development of the egg into a tadpole. However, he found that the potential of a transplanted nucleus to direct normal development was inversely related to the age of the donor: The older the donor nucleus, the lower the percentage of normal tadpoles (see Figure 20.16).

The other major source of developmental information, which becomes increasingly important as the number of embryonic cells increases, is the environment around a particular cell. Most influential are the signals conveyed to an embryonic cell from other embryonic cells in the vicinity, including contact with cell-surface molecules on neighboring cells and the binding of growth factors secreted by neighboring cells (see Concept 11.1).

Such signals cause changes in the target cells, a process called induction (Figure 18.17b). The molecules passing along these signals within the target cell are cell-surface receptors and other signaling pathway proteins. In general, the signaling molecules send a cell down a specific developmental path by causing changes in its gene expression that eventually result in observable cellular changes. Thus, interactions between embryonic cells help induce differentiation into the many specialized cell types making up a new organism.

A single ribosome can make an average-sized polypeptide in less than a minute. In both bacteria and eukaryotes, however, multiple ribosomes translate an mRNA at the same time (Figure 17.23); that is, a single mRNA is used to make many copies of a polypeptide simultaneously. Once a ribosome is far enough past the start codon, a second ribosome can attach to the mRNA, eventually resulting in a number of ribosomes trailing along the mRNA.

Such strings of ribosomes, called polyribosomes (or polysomes), can be seen with an electron microscope; they can be either free or bound. They enable a cell to rapidly make many copies of a polypeptide. Another way both bacteria and eukaryotes augment the number of copies of a polypeptide is by transcribing multiple mRNAs from the same gene. However, the coordination of the two processes—transcription and translation—differs in the two groups. The most important differences between bacteria and eukaryotes arise from the bacterial cell's lack of compartmental organization. Like a one-room workshop, a bacterial cell ensures a streamlined operation by coupling the two processes. In the absence of a nucleus, it can simultaneously transcribe and translate the same gene (Figure 17.24), and the newly made protein can quickly diffuse to its site of function.

For the present, the diagrams in Figure 18.24 and Figure 18.25 are an accurate view of how mutations can contribute to cancer, but we still don't know exactly how a particular cell becomes a cancer cell. As we discover previously unknown aspects of gene regulation, it is informative to study their role in the onset of cancer.

Such studies have shown, for instance, that DNA methylation and histone modification patterns differ in normal and cancer cells and that miRNAs probably participate in cancer development. While we've learned a lot about cancer by studying cell-signaling pathways, there is still much more to learn.

Given a genotypic ratio of 1 AA:2 Aa :1 aa for offspring of an Aa * Aa cross, John and Carol each have a 2 ⁄ 3 chance of being carriers (Aa). According to the rule of multiplication, the overall probability of their firstborn having the disorder is 2 ⁄ 3 (the chance that John is a carrier) times 2 ⁄ 3 (the chance that Carol is a carrier) times 1 ⁄ 4 (the chance of two carriers having a child with the disease), which equals 1 ⁄ 9.

Suppose that Carol and John decide to have a child—after all, there is an 8 ⁄ 9 chance that their baby will not have the disorder. If, despite these odds, their child is born with the disease, then we would know that both John and Carol are, in fact, carriers (Aa genotype). If both John and Carol are carriers, there is ⁄ 4 chance that any subsequent child this couple has will have the disease. The probability is higher for subsequent children because the diagnosis of the disease in the first child established that both parents are carriers, not because the genotype of the first child affects in any way that of future children.

A closer look at the relationship between dominance and phenotype reveals an intriguing fact: For any character, the observed dominant/recessive relationship of alleles depends on the level at which we examine phenotype. Tay-Sachs disease, an inherited disorder in humans, is an example

The brain cells of a child with Tay-Sachs disease cannot metabolize certain lipids because a crucial enzyme does not work properly. As these lipids accumulate in brain cells, the child begins to suffer seizures, blindness, and degeneration of motor and mental performance and dies within a few years.

A linkage map does portray the order of genes along a chromosome, but it does not accurately portray the precise locations of those genes. Other methods enable geneticists to construct cytogenetic maps of chromosomes, which locate genes with respect to chromosomal features, such as stained bands, that can be seen in the microscope.

Technical advances over the last two decades have enormously increased the rate and affordability of DNA sequencing. Therefore, today, most researchers sequence whole genomes to map the locations of genes of a given species. The entire nucleotide sequence is the ultimate physical map of a chromosome, revealing the physical distances in DNA nucleotides between gene loci (see Concept 21.1). Comparing a linkage map with such a physical map or with a cytogenetic map of the same chromosome, we find that the linear order of genes is identical in all the maps, but the spacing between genes is not.

Research Method :Crossing Pea Plants Application: By crossing (mating) two true-breeding varieties of an organism, scientists can study patterns of inheritance. In this example, Mendel crossed pea plants that varied in flower color.

Technique: Parental generation (P): (1) Removed stamens from purple flower (2) Transferred spermbearing pollen from stamens of white flower to eggbearing carpel of purple flower (3) Waited for pollinated carpel to mature into pod. (4) Planted seeds from pod 4. Results : When pollen from a white flower was transferred to a purple flower, the first-generation hybrids all had purple flowers. The result was the same for the reciprocal cross, which involved the transfer of pollen from purple flowers to white flowers. First filial generation offspring (F1) (5) Examined offspring: all purple flowers

Hundreds of different restriction enzymes have been identified and isolated. Each restriction enzyme is very specific, recognizing a particular short DNA sequence, or restriction site, and cutting both DNA strands at precise points within this restriction site.

The DNA of a bacterial cell is protected from the cell's own restriction enzymes by the addition of methyl groups (¬CH3) to adenines or cytosines within the sequences recognized by the enzymes.

Imagine crossing two true-breeding pea varieties that differ in both of these characters—a cross between a plant with yellow-round seeds (YYRR) and a plant with green-wrinkled seeds (yyrr).

The F1 plants will be dihybrids, individuals heterozygous for the two characters being followed in the cross (YyRr). But are these two characters transmitted from parents to offspring as a package? That is, will the Y and R alleles always stay together, generation after generation? Or are seed color and seed shape inherited independently? Figure 14.8 shows how a dihybrid cross, a cross between F1 dihybrids, can determine which of these two hypotheses is correct.

These tests for identifying carriers enable people with family histories of genetic disorders to make informed decisions about having children, including whether to do genetic testing of the fetus, should they decide to become pregnant. The tests also raise other issues: Could carriers be denied health or life insurance or lose the jobs providing those benefits, even though they themselves are healthy?

The Genetic Information Nondiscrimination Act, signed into law in the United States in 2008, allays these concerns by prohibiting discrimination in employment or insurance coverage based on genetic test results. A question that remains is whether sufficient genetic counseling is available to help large numbers of individuals understand their genetic test results. Even when test results are clearly understood, affected individuals may still face difficult decisions. Advances in biotechnology offer the potential to reduce human suffering, but along with them come ethical issues that require conscientious deliberation.

Replicative Cycles of Phages: Phages are the best understood of all viruses, although some of them are also among the most complex. Research on phages led to the discovery that some double-stranded DNA viruses can replicate by two alternative mechanisms: the lytic cycle and the lysogenic cycle.

The Lytic Cycle: A phage replicative cycle that culminates in death of the host cell is known as a lytic cycle. The term refers to the last stage of infection, during which the bacterium lyses (breaks open) and releases the phages that were produced within the cell. Each of these phages can then infect a healthy cell, and a few successive lytic cycles can destroy an entire bacterial population in just a few hours. A phage that replicates only by a lytic cycle is a virulent phage. Figure 19.5 illustrates the major steps in the lytic cycle of T4, a typical virulent phage.

The HIV replicative cycle (traced in Figure 19.9) is typical of a retrovirus. After HIV enters a host cell, its reverse transcriptase molecules are released into the cytoplasm, where they catalyze synthesis of viral DNA. The newly made viral DNA then enters the cell's nucleus and integrates into the DNA of a chromosome. The integrated viral DNA, called a provirus, never leaves the host's genome, remaining a permanent resident of the cell. (Recall that a prophage, in contrast, leaves the host's genome at the start of a lytic cycle.)

The RNA polymerase of the host transcribes the proviral DNA into RNA molecules, which can function both as mRNA for the synthesis of viral proteins and as genomes for the new viruses that will be assembled and released from the cell. In Concept 43.4, we describe how HIV causes the deterioration of the immune system that occurs in AIDS.

Furthermore, sexual reproduction is more expensive energetically than asexual reproduction. In spite of these apparent disadvantages, sexual reproduction is almost universal among animals. Why is this?

The ability of sexual reproduction to generate genetic diversity is the most commonly proposed explanation for the evolutionary persistence of this process. However, consider the unusual case of the bdelloid rotifer (Figure 13.13).

n addition to regulating the lac operon, CRP helps regulate other operons that encode enzymes used in catabolic pathways. All told, it may affect the expression of more than 100 genes in E. coli. When glucose is plentiful and CRP is inactive, the synthesis of enzymes that catabolize compounds other than glucose generally slows down.

The ability to catabolize other compounds, such as lactose, enables a cell deprived of glucose to survive. The compounds present in any given cell at the moment determine which operons are switched on—the result of simple interactions of activator and repressor proteins with the promoters of the genes in question.

Cross-Species Gene Expression and Evolutionary Ancestry:

The ability to express eukaryotic proteins in bacteria (even if the proteins aren't modified properly) is quite remarkable when we consider how different eukaryotic and bacterial cells are. In fact, examples abound of genes that are taken from one species and function perfectly well when transferred into another very different species, such as a firefly gene in a tobacco plant and a jellyfish gene in a pig (see Figure 17.7). These observations underscore the shared evolutionary ancestry of species living today.

Today, we know of countless examples in which a mutation in a gene causes a faulty enzyme that in turn leads to an identifiable condition. The albino donkey in Figure 17.1 lacks a key enzyme called tyrosinase in the metabolic pathway that produces melanin, a dark pigment.

The absence of melanin causes white fur and other effects throughout the donkey's body. Its nose, ears, and hooves, as well as its eyes, are pink because no melanin is present to mask the reddish color of the blood vessels that run through those structures.

The accurate translation of a genetic message requires two instances of molecular recognition. First, a tRNA that binds to an mRNA codon specifying a particular amino acid must carry that amino acid, and no other, to the ribosome. The correct matching up of tRNA and amino acid is carried out by a family of related enzymes that are aptly named aminoacyl-tRNA synthetases (Figure 17.17).

The active site of each type of aminoacyl-tRNA synthetase fits only a specific combination of amino acid and tRNA. There are 20 different synthetases, one for each amino acid. A synthetase joins a given amino acid to an appropriate tRNA; one synthetase is able to bind to all the different tRNAs for its particular amino acid. The synthetase catalyzes the covalent attachment of the amino acid to its tRNA in a process driven by the hydrolysis of ATP. The resulting aminoacyl tRNA, also called a charged tRNA, is released from the enzyme and is then available to deliver its amino acid to a growing polypeptide chain on a ribosome.

A metabolic pathway can be controlled on two levels, as shown for the synthesis of tryptophan in Figure 18.2. First, cells can adjust the activity of enzymes already present. This is a fairly rapid response, which relies on the sensitivity of many enzymes to chemical cues that increase or decrease their catalytic activity (see Concept 8.5).

The activity of the first enzyme in the pathway is inhibited by the pathway's end product— tryptophan, in this case (Figure 18.2a). Thus, if tryptophan accumulates in a cell, it shuts down the synthesis of more tryptophan by inhibiting enzyme activity. Such feedback inhibition, typical of anabolic (biosynthetic) pathways, allows a cell to adapt to short-term fluctuations in the supply of a substance it needs (see Figure 8.21).

Antiparallel Elongation: As we have noted previously, the two ends of a DNA strand are different, giving each strand directionality, like a one-way street (see Figure 16.5). In addition, the two strands of DNA in a double helix are antiparallel, meaning that they are oriented in opposite directions to each other, like the two sides of a divided street (see Figure 16.14). Therefore, the two new strands formed during DNA replication must also be antiparallel to their template strands.

The antiparallel arrangement of the double helix, together with a property of DNA polymerases, has an important effect on how replication occurs. Because of their structure, DNA polymerases can add nucleotides only to the free 3′ end of a primer or growing DNA strand, never to the 5′ end (see Figure 16.14). Thus, a new DNA strand can elongate only in the 5′ S 3′ direction. With this in mind, let's examine one of the two replication forks in a bubble (Figure 16.15). Along one template strand, DNA polymerase III can synthesize a complementary strand continuously by elongating the new DNA in the mandatory 5′ S 3′ direction. DNA pol III remains in the replication fork on that template strand and continuously adds nucleotides to the new complementary strand as the fork progresses. The DNA strand made by this mechanism is called the leading strand. Only one primer is required for DNA pol III to synthesize the entire leading strand (see Figure 16.15).

The most useful restriction enzymes cleave the sugarphosphate backbones in the two DNA strands in a staggered manner, as indicated in Figure 20.5. The resulting doublestranded restriction fragments have at least one single-stranded end, called a sticky end. These short extensions can form hydrogen-bonded base pairs with complementary sticky ends on any other DNA molecules cut with the same enzyme.

The associations formed in this way are only temporary but can be made permanent by DNA ligase, which catalyzes the formation of covalent bonds that close up the sugar-phosphate backbones of DNA strands (see Figure 16.16). You can see at the bottom of Figure 20.5 that the ligase-catalyzed joining of DNA from two different sources produces a stable recombinant DNA molecule, in this example, a recombinant plasmid.

Basal-like: ERα- • PR- • HER2- • 15-20% of breast cancers • More aggressive; poorer prognosis than other subtypes

The basal-like subtype is "triple negative"—it does not express ERα, PR, or HER2. It often has a mutation in the tumor-suppressor gene BRCA1 (see Concept 18.5). Treatments that target ER, PR, or HER2 are not effective, but new treatments are being developed. Currently, patients are treated with cytotoxic chemotherapy, which selectively kills fast-growing cells.

Thus, the first components to associate with each other during the initiation stage of translation are mRNA, a tRNA bearing the first amino acid of the polypeptide, and the small ribosomal subunit (Figure 17.19). This is followed by the attachment of a large ribosomal subunit, completing the translation initiation complex. Proteins called initiation factors are required to bring all these components together.

The cell also expends energy obtained by hydrolysis of a GTP molecule to form the initiation complex. At the completion of the initiation process, the initiator tRNA sits in the P site of the ribosome, and the vacant A site is ready for the next aminoacyl tRNA. Note that a polypeptide is always synthesized in one direction, from the initial methionine at the amino end, also called the N-terminus, toward the final amino acid at the carboxyl end, also called the C-terminus (see Figure 5.15).

In an alternative technique called chorionic villus sampling (CVS), a physician inserts a narrow tube through the cervix into the uterus and suctions out a tiny sample of tissue from the placenta, the organ that transmits nutrients and fetal wastes between the fetus and the mother (Figure 14.19b).

The cells of the chorionic villi of the placenta—the portion sampled—are derived from the fetus and have the same genotype and DNA sequence as the new individual. These cells are proliferating rapidly enough to allow karyotyping to be carried out immediately. This rapid analysis represents an advantage over amniocentesis, in which the cells must be cultured for several weeks before karyotyping. Another advantage of CVS is that it can be performed as early as the 10th week of pregnancy.

If a point mutation occurs in a gamete or in a cell that gives rise to gametes, it may be transmitted to offspring and to future generations. If the mutation has an adverse effect on the phenotype of a person, the mutant condition is referred to as a genetic disorder or hereditary disease. For example, we can trace the genetic basis of sickle-cell disease to the mutation of a single nucleotide pair in the gene that encodes the β-globin polypeptide of hemoglobin.

The change of a single nucleotide in the DNA's template strand leads to an altered mRNA and the production of an abnormal protein (Figure 17.26; also see Figure 5.19). In individuals who are homozygous for the mutant allele, the sickling of red blood cells caused by the altered hemoglobin produces the multiple symptoms associated with sickle-cell disease (see Concept 14.4 and Figure 23.18). Another disorder caused by a point mutation is a heart condition called familial cardiomyopathy, which is responsible for some of the tragic incidents of sudden death in young athletes. Point mutations in several genes encoding muscle proteins have been identified, any of which can lead to this disorder.

Each sexually reproducing species has a characteristic diploid and haploid number. For example, the fruit fly Drosophila melanogaster has a diploid number (2n) of 8 and a haploid number (n) of 4, while for dogs, 2n is 78 and n is 39

The chromosome number generally does not correlate with the size or complexity of a species' genome; it simply reflects how many linear pieces of DNA make up the genome, which is a function of the evolutionary history of that species (see Concept 21.5). Now let's consider chromosome behavior during sexual life cycles. We'll use the human life cycle as an example.

A stem cell is a relatively unspecialized cell that can both reproduce itself indefinitely and, under appropriate conditions, differentiate into specialized cells of one or more types. Stem cells have great potential for regenerating damaged tissues.

The cloning of plants and animals was first attempted over 50 years ago in experiments designed to answer basic biological questions. For example, researchers wondered if all the cells of an organism have the same genes or whether cells lose genes during the process of differentiation (see Concept 18.4). One way to answer this question is to see whether a differentiated cell can generate a whole organism—in other words, whether cloning an organism is possible. Let's discuss these early experiments before we consider more recent progress in organismal cloning and procedures for producing stem cells.

Studying the Expression of Single Genes: Suppose we have cloned a gene that we suspect plays an important role in the embryonic development of Drosophila melanogaster (the fruit fly). The first thing we might want to know is which embryonic cells express the gene—in other words, where in the embryo is the corresponding mRNA found? We can detect the mRNA by nucleic acid hybridization with molecules of complementary sequence that we can follow in some way.

The complementary molecule, a short, single-stranded nucleic acid that can be either RNA or DNA, is called a nucleic acid probe. Using our cloned gene as a template, we can synthesize a probe complementary to the mRNA. For example, if part of the sequence on the mRNA were 5′ ...CUCAUCACCGGC ... 3′ then we would synthesize this single-stranded DNA probe: 3′ GAGTAGTGGCCG 5′ Each probe molecule is labeled during synthesis with a fluorescent tag so we can follow it. A solution containing probe molecules is applied to Drosophila embryos, allowing the probe to hybridize specifically with any complementary sequences on the many mRNAs in embryonic cells in which the gene is being transcribed. . Because this technique allows us to see the mRNA in place (or in situ, in Latin) in the intact organism, this technique is called in situ hybridization. Different probes can be labeled with different fluorescent dyes, sometimes with strikingly beautiful results (Figure 20.9).

Evidence suggests that this foreign DNA can become incorporated into the genome of the bdelloid, leading to increased genetic diversity. In fact, the genomic analysis shows that bdelloid rotifers pick up nonbdelloid DNA at a much higher rate than most other species pick up foreign DNA.

The conclusion that bdelloid rotifers have developed other ways of achieving genetic diversity supports the idea that genetic diversity is advantageous, but that sexual reproduction is not the only way of generating such diversity.

A single gene can be transcribed simultaneously by several molecules of RNA polymerase following each other like trucks in a convoy. A growing strand of RNA trails off from each polymerase, with the length of each new strand reflecting how far along the template the enzyme has traveled from the start point (see the mRNA molecules in Figure 17.23).

The congregation of many polymerase molecules simultaneously transcribing a single gene increases the amount of mRNA transcribed from it, which helps the cell make the encoded protein in large amounts.

Emerging Viruses: Viruses that suddenly become apparent are often referred to as emerging viruses. HIV, the AIDS virus, is a classic example: This virus appeared in San Francisco in the early 1980s, seemingly out of nowhere, although later studies uncovered a case in the Belgian Congo in 1959. A number of other dangerous emerging viruses cause encephalitis, inflammation of the brain. One example is the West Nile virus, which appeared in North America in 1999 and has spread to all 48 contiguous states in the United States, by now resulting in over 40,000 cases and almost 2,000 deaths

The deadly Ebola virus (Figure 19.10a), recognized initially in 1976 in central Africa, is one of several emerging viruses that cause hemorrhagic fever, an often fatal illness characterized by fever, vomiting, massive bleeding, and circulatory system collapse. In 2014, a widespread outbreak of Ebola virus in western Africa caused the World Health Organization to declare an international health emergency. By mid-2015 the outbreak, centered in Guinea, Sierra Leone, and Liberia, had caused over 27,000 illnesses and 11,000 deaths.

As you learned in Figure 13.9, crossing over produces chromosomes with new combinations of maternal and paternal alleles. At metaphase II, chromosomes that contain one or more recombinant chromatids can be oriented in two alternative, nonequivalent ways with respect to other chromosomes because their sister chromatids are no longer identical (see Figure 13.12).

The different possible arrangements of nonidentical sister chromatids during meiosis II further increase the number of genetic types of daughter cells that can result from meiosis. You'll learn more about crossing over in Chapter 15. The important point for now is that crossing over, by combining DNA inherited from two parents into a single chromosome, is an important source of genetic variation in sexual life cycles.

Mapping the Distance Between Genes Using Recombination Data: Scientific Inquiry

The discovery of linked genes and recombination due to crossing over motivated one of Morgan's students, Alfred H. Sturtevant, to work out a method for constructing a genetic map, an ordered list of the genetic loci along a particular chromosome.

DNA sequencing and DNA cloning are valuable tools for genetic engineering and biological inquiry:

The discovery of the structure of the DNA molecule, and specifically the recognition that its two strands are complementary to each other, opened the door for the development of DNA sequencing and other techniques used in biological research today. Key to these techniques is nucleic acid hybridization, the base pairing of one strand of a nucleic acid to a complementary sequence on a strand from a different nucleic acid molecule. In this section, we'll first describe DNA-sequencing techniques. Then we'll explore other important methods used in genetic engineering, the direct manipulation of genes for practical purposes

Another class of small RNAs, similar in size and function to miRNAs, is called small interfering RNAs (siRNAs). Both miRNAs and siRNAs can associate with the same proteins, producing similar results. In fact, if siRNA precursor RNA molecules are injected into a cell, the cell's machinery can process them into siRNAs that turn off expression of genes with related sequences, similarly to how miRNAs function.

The distinction between miRNAs and siRNAs is based on subtle differences in the structure of their precursors, which in both cases are RNA molecules that are mostly double-stranded.

The mRNA is moved through the ribosome in one direction only, 5' end first; this is equivalent to the ribosome moving 5' -> 3' on the mRNA. The main point is that the ribosome and the mRNA move relative to each other, unidirectionally, codon by codon.

The elongation cycle takes less than a tenth of a second in bacteria and is repeated as each amino acid is added until the polypeptide is complete. The empty tRNAs that are released from the E site return to the cytoplasm, where they will be reloaded with the appropriate amino acid (see Figure 17.17).

Most cellular systems for repairing incorrectly paired nucleotides, whether they are due to DNA damage or to replication errors, use a mechanism that takes advantage of the base-paired structure of DNA. In many cases, a segment of the strand containing the damage is cut out (excised) by a DNA cutting enzyme—a nuclease—and the resulting gap is then filled in with nucleotides, using the undamaged strand as a template.

The enzymes involved in filling the gap are a DNA polymerase and DNA ligase. One such DNA repair system is called nucleotide excision repair (Figure 16.19).

The crowd at a soccer match attests to the marvelous variety and diversity of humankind. Brown, blue, or gray eyes; black, brown, or blond hair—these are just a few examples of heritable variations that we may observe. What principles account for the transmission of such traits from parents to offspring?

The explanation of heredity most widely in favor during the 1800s was the "blending" hypothesis, the idea that genetic material contributed by the two parents mixes just as blue and yellow paints blend to make green. This hypothesis predicts that over many generations, a freely mating population will give rise to a uniform population of individuals, something we don't see. The blending hypothesis also fails to explain how traits can reappear after they've skipped a generation.

In Figure 18.11, you saw a simplified view of how differential gene expression occurs in two cell types, a liver cell and a lens cell. Each of these fully differentiated cells has a particular mix of specific activators that turn on the collection of genes whose products are required in the cell.

The fact that both cells arose through a series of mitoses from a common fertilized egg inevitably leads to a question: How do different sets of activators come to be present in the two cells? It turns out that materials placed into the egg by maternal cells set up a sequential program of gene regulation that is carried out as embryonic cells divide, and this program coordinates cell differentiation during embryonic development. To understand how this works, we will consider two basic developmental processes. First, we'll explore how cells that arise from early embryonic mitoses develop the differences that start each cell along its own differentiation pathway. Second, we'll see how cellular differentiation leads to one particular cell type, using muscle development as an example.

Inherited Predisposition and Environmental Factors Contributing to Cancer:

The fact that multiple genetic changes are required to produce a cancer cell helps explain the observation that cancers can run in families. An individual inheriting an oncogene or a mutant allele of a tumor-suppressor gene is one step closer to accumulating the necessary mutations for cancer to develop than is an individual without any such mutations

The DNA Toolbox: The last decade or so has seen some extraordinary feats in biology, among them determination of the complete DNA sequences of several extinct species, including woolly mammoths (see below), Neanderthals, and a 700,000-year-old horse. Pivotal to those discoveries was the sequencing of the human genome, essentially completed in 2003. This endeavor marked a turning point in biology because it sparked remarkable technological advances in DNA sequencing

The first human genome sequence took several years at a cost of 1 billion dollars; the time and cost of sequencing a genome have been in free fall since then. Figure 20.1 shows a model of a sequencing technique in which the nucleotides of a single strand of DNA are passed one by one through a very small pore in a membrane, and the resulting tiny changes in an electrical current are used to determine the nucleotide sequence. Developers of this technique, which you will learn more about later in the chapter, claim that ultimately we will be able to sequence a human genome in about 6 hours on a $900 device the size of a pack of gum.

The model of a multistep path to cancer is well supported by studies of one of the best-understood types of human cancer: colorectal cancer, which affects the colon and/or rectum. About 140,000 new cases of colorectal cancer are diagnosed each year in the United States, and the disease causes 50,000 deaths per year. Like most cancers, colorectal cancer develops gradually (Figure 18.26).

The first sign is often a polyp, a small, benign growth in the colon lining. The cells of the polyp look normal, although they divide unusually frequently. The tumor grows and may eventually become malignant, invading other tissues. The development of a malignant tumor is paralleled by a gradual accumulation of mutations that convert proto-oncogenes to oncogenes and knock out tumor-suppressor genes. A ras oncogene and a mutated p53 tumor-suppressor gene are often involved.

Morgan showed that Mendelian inheritance has its physical basis in the behavior of chromosomes: scientific inquiry

The first solid evidence associating a specific gene with a specific chromosome came early in the 1900s from the work of Thomas Hunt Morgan, an experimental embryologist at Columbia University. Although Morgan was initially skeptical about both Mendelian genetics and the chromosome theory, his early experiments provided convincing evidence that chromosomes are indeed the location of Mendel's heritable factors.

In Figure 20.4, the plasmid acts as a cloning vector, a DNA molecule that can carry foreign DNA into a host cell and be replicated there. Bacterial plasmids are widely used as cloning vectors for several reasons: They can be readily obtained from commercial suppliers, manipulated to form recombinant plasmids by insertion of foreign DNA in a test tube (referred to as in vitro, from the Latin meaning "in glass"), and then easily introduced into bacterial cells.

The foreign DNA in Figure 20.4 is a gene from a eukaryotic cell; we will describe in more detail how the foreign DNA segment was obtained later in this section.

Random Fertilization: The random nature of fertilization adds to the genetic variation arising from meiosis. In humans, each male and female gamete represents one of about 8.4 million (2^23) possible chromosome combinations due to independent assortment.

The fusion of a male gamete with a female gamete during fertilization will produce a zygote with any of about 70 trillion (2^23 * 2^23) diploid combinations. If we factor in the variation brought about by crossing over, the number of possibilities is truly astronomical. It may sound trite, but you really are unique.

Cancer results from genetic changes that affect cell cycle control: In Concept 12.3, we considered cancer as a type of disease in which cells escape from the control mechanisms that normally limit their growth. Now that we have discussed the molecular basis of gene expression and its regulation, we are ready to look at cancer more closely.

The gene regulation systems that go wrong during cancer turn out to be the very same systems that play important roles in embryonic development, the immune response, and many other biological processes. Thus, research into the molecular basis of cancer has both benefited from and informed many other fields of biology

Triplets of nucleotide bases are the smallest units of uniform length that can code for all the amino acids. If each arrangement of three consecutive nucleotide bases specifies an amino acid, there can be 64 (that is, 43 ) possible code words—more than enough to specify all the amino acids. Experiments have verified that the flow of information from gene to protein is based on a triplet code:

The genetic instructions for a polypeptide chain are written in the DNA as a series of nonoverlapping, three-nucleotide words. The series of words in a gene is transcribed into a complementary series of nonoverlapping, three-nucleotide words in mRNA, which is then translated into a chain of amino acids (Figure 17.5).

General Features of Viral Replicative Cycles: A viral infection begins when a virus binds to a host cell and the viral genome makes its way inside (Figure 19.4). The mechanism of genome entry depends on the type of virus and the type of host cell. For example, T-even phages use their elaborate tail apparatus to inject DNA into a bacterium (see Figure 19.3d). Other viruses are taken up by endocytosis or, in the case of enveloped viruses, by fusion of the viral envelope with the host's plasma membrane. Once the viral genome is inside, the proteins it encodes can commandeer the host, reprogramming the cell to copy the viral genome and manufacture viral proteins.

The host provides the nucleotides for making viral nucleic acids, as well as enzymes, ribosomes, tRNAs, amino acids, ATP, and other components needed for making the viral proteins. Many DNA viruses use the DNA polymerases of the host cell to synthesize new genomes along the templates provided by the viral DNA. In contrast, to replicate their genomes, RNA viruses use virally encoded RNA polymerases that can use RNA as a template. (Uninfected cells generally make no enzymes for carrying out this process.)

Behavior of Chromosome Sets in the Human Life Cycle:

The human life cycle begins when a haploid sperm from the father fuses with a haploid egg from the mother (Figure 13.5).This union of gametes, culminating in fusion of their nuclei, is called fertilization. The resulting fertilized egg, or zygote, is diploid because it contains two haploid sets of chromosomes bearing genes representing the maternal and paternal family lines.

sorders are treatable at present. Fetal and newborn screening for serious inherited diseases, tests for identifying carriers, and genetic counseling all rely on the Mendelian model of inheritance. We owe the "gene idea"—the concept of heritable factors transmitted according to simple rules of chance—to the elegant quantitative experiments of Gregor Mendel.

The importance of his discoveries was overlooked by most biologists until early in the 20th century, decades after he reported his findings. In the next chapter, you will learn how Mendel's laws have their physical basis in the behavior of chromosomes during sexual life cycles and how the synthesis of Mendelian genetics and a chromosome theory of inheritance catalyzed progress in genetics.

X Inactivation in Female Mammals: Female mammals, including human females, inherit two X chromosomes—twice the number inherited by males—so you may wonder whether females make twice as many of the proteins encoded by X-linked genes as males. In fact, almost all of one X chromosome in each cell in female mammals becomes inactivated during early embryonic development. As a result, the cells of females and males have the same effective dose (one active copy) of most X-linked genes.

The inactive X in each cell of a female condenses into a compact object called a Barr body (discovered by Canadian anatomist Murray Barr), which lies along the inside of the nuclear envelope. Most of the genes of the X chromosome that forms the Barr body are not expressed. In the ovaries, however, Barr body chromosomes are reactivated in the cells that give rise to eggs, resulting in every female gamete (egg) having an active X after meiosis

Extending Mendelian Genetics for a Single Gene:

The inheritance of characters determined by a single gene deviates from simple Mendelian patterns when alleles are not completely dominant or recessive, when a particular gene has more than two alleles, or when a single gene produces multiple phenotypes. We will describe examples of each of these situations in this section.

. Proteinprotein interactions are crucial to the initiation of eukaryotic transcription. Only when the complete initiation complex has assembled can the polymerase begin to move along the DNA template strand, producing a complementary strand of RNA.

The interaction of general transcription factors and RNA polymerase II with a promoter usually leads to a low rate of initiation and production of few RNA transcripts from genes that are not expressed all the time, but instead are regulated. In eukaryotes, high levels of transcription of these particular genes at the appropriate time and place depend on the interaction of control elements with another set of proteins, which can be thought of as specific transcription factors.

How is pre-mRNA splicing carried out? The removal of introns is accomplished by a large complex made of proteins and small RNAs called a spliceosome. This complex binds to several short nucleotide sequences along an intron, including key sequences at each end (Figure 17.13).

The intron is then released (and rapidly degraded), and the spliceosome joins together the two exons that flanked the intron. It turns out that the small RNAs in the spliceosome not only participate in spliceosome assembly and splice site recognition, but also catalyze the splicing reaction.

Regulation of Chromatin Structure: Recall that the DNA of eukaryotic cells is packaged with proteins in an elaborate complex known as chromatin, the basic unit of which is the nucleosome (see Figure 16.22). The structural organization of chromatin not only packs a cell's DNA into a compact form that fits inside the nucleus, but also helps regulate gene expression in several ways.

The location of a gene's promoter, relative to both placement of nucleosomes and the sites where the DNA attaches to the chromosome scaffold, can affect whether the gene is transcribed. In addition, genes within heterochromatin, which is highly condensed, are usually not expressed. Lastly, certain chemical modifications to chromatin—both to the histone proteins of the nucleosomes around which DNA is wrapped and to the nucleotides that make up that DNA—can influence chromatin structure and gene expression. Here we examine the effects of these modifications, which are catalyzed by specific enzymes.

What mechanisms generate this genetic variation?

The key is the behavior of chromosomes during the sexual life cycle

A Mendelian View of Heredity and Variation: We have now broadened our view of Mendelian inheritance by exploring degrees of dominance as well as multiple alleles, pleiotropy, epistasis, polygenic inheritance, and the phenotypic impact of the environment. How can we integrate these refinements into a comprehensive theory of Mendelian genetics?

The key is to make the transition from the reductionist emphasis on single genes and phenotypic characters to the emergent properties of the organism as a whole, one of the themes of this book.

The Structure and Function of Transfer RNA:

The key to translating a genetic message into a specific amino acid sequence is the fact that each tRNA molecule enables translation of a given mRNA codon into a certain amino acid. This is possible because a tRNA bears a specific amino acid at one end of its three-dimensional structure, while at the other end is a nucleotide triplet that can base-pair with the complementary codon on mRNA.

Thus, sister chromatid cohesion and crossing over, acting together, play an essential role in the lining up of chromosomes by homologous pairs at metaphase I.

The molecular basis of chromosome behavior during meiosis continues to be a focus of intense research. In the Scientific Skills Exercise, you can work with data tracking the amount of DNA in cells as they progress through meiosis.

The fish in Figure 18.1 is keeping an eye out for predators—or, more precisely, both halves of each eye! Anableps anableps, commonly known as "cuatro ojos" ("four eyes"), glides through freshwater lakes and ponds in Central and South America with the upper half of each eye protruding from the water. The eye's upper half is particularly well-suited for aerial vision and the lower half for aquatic vision.

The molecular basis of this specialization has recently been revealed: The cells of the two parts of the eye express a slightly different set of genes involved in vision, even though these two groups of cells are quite similar and contain identical genomes. What is the biological mechanism underlying the difference in gene expression that makes this remarkable feat possible?

Figure 20.5 shows how restriction enzymes are used to clone a foreign DNA fragment into a bacterial plasmid. At the top is a bacterial plasmid (like the one in Figure 20.4) that has a single restriction site recognized by a particular restriction enzyme from E. coli. As shown in this example, most restriction sites are symmetrical. That is, the sequence of nucleotides is the same on both strands when read in the 5¿ S 3¿ direction.

The most commonly used restriction enzymes recognize sequences containing four to eight nucleotide pairs. Because any sequence this short usually occurs (by chance) many times in a long DNA molecule, a restriction enzyme will make many cuts in such a DNA molecule, yielding a set of restriction fragments. All copies of a given DNA molecule always yield the same set of restriction fragments when exposed to the same restriction enzyme.

Plants and some species of algae exhibit a second type of life cycle called alternation of generations (Figure 13.6b). This type includes both diploid and haploid stages that are multicellular.

The multicellular diploid stage is called the sporophyte. Meiosis in the sporophyte produces haploid cells called spores.

How Linkage Affects Inheritance: To see how linkage between genes affects the inheritance of two different characters, let's examine another of Morgan's Drosophila experiments. In this case, the characters are body color and wing size, each with two different phenotypes. Wildtype flies have gray bodies and normal-sized wings. In addition to these flies, Morgan had managed to obtain, through breeding, doubly mutant flies with black bodies and wings much smaller than normal, called vestigial wings.

The mutant alleles are recessive to the wild-type alleles, and neither gene is on a sex chromosome. In his investigation of these two genes, Morgan carried out the crosses shown in Figure 15.9. The first was a P generation cross to generate F1 dihybrid flies, and the second was a testcross.

Cancer research led to the discovery of cancer-causing genes called oncogenes (from the Greek onco, tumor) in certain types of viruses. Subsequently, close counterparts of viral oncogenes were found in the genomes of humans and other animals

The normal versions of the cellular genes, called proto-oncogenes, code for proteins that stimulate normal cell growth and division.

Linked genes tend to be inherited together because they are located near each other on the same chromosome:

The number of genes in a cell is far greater than the number of chromosomes; in fact, each chromosome (except the Y) has hundreds or thousands of genes. Genes located near each other on the same chromosome tend to be inherited together in genetic crosses; such genes are said to be genetically linked and are called linked genes. When geneticists follow linked genes in breeding experiments, the results deviate from those expected from Mendel's law of independent assortment.

Evolutionary Developmental Biology ("Evo-Devo") The fly with legs emerging from its head in Figure 18.20 is the result of a single mutation in one gene, a homeotic gene. The gene does not encode any antenna protein, however. Instead, it encodes a transcription factor that regulates other genes, and its malfunction leads to misplaced structures, such as legs instead of antennae.

The observation that a change in gene regulation during development could lead to such a fantastic change in body form prompted some scientists to consider whether these types of mutations could contribute to evolution by generating novel body shapes. Ultimately, this line of inquiry gave rise to the field of evolutionary developmental biology, so-called "evo-devo," which will be further discussed in Concept 21.6.

A key advantage of grouping genes of related function into one transcription unit is that a single "on-off switch" can control the whole cluster of functionally related genes; in other words, these genes are coordinately controlled. When an E. coli cell must make tryptophan for itself because its surrounding environment lacks this amino acid, all the enzymes for the metabolic pathway are synthesized at the same time.

The on-off switch is a segment of DNA called an operator. Both its location and name suit its function: Positioned within the promoter or, in some cases, between the promoter and the enzyme-coding genes, the operator controls the access of RNA polymerase to the genes. Together, the operator, the promoter, and the genes they control—the entire stretch of DNA required for enzyme production for the tryptophan pathway—constitute an operon. The trp operon (trp for tryptophan) is one of many operons in the E. coli genome (see Figure 18.3a).

Herbivores, especially insects, pose a double threat because they can also act as carriers of viruses, transmitting disease from plant to plant. Moreover, farmers and gardeners may transmit plant viruses inadvertently on pruning shears and other tools.

The other route of viral infection is vertical transmission, in which a plant inherits a viral infection from a parent. Vertical transmission can occur in asexual propagation (for example, through cuttings) or in sexual reproduction via infected seeds.

Once a virus enters a plant cell and begins replicating, viral genomes and associated proteins can spread throughout the plant by means of plasmodesmata, the cytoplasmic connections that penetrate the walls between adjacent plant cells (see Figure 36.19).

The passage of viral macromolecules from cell to cell is facilitated by virally encoded proteins that cause enlargement of plasmodesmata. Scientists have not yet devised cures for most viral plant diseases. Consequently, research efforts are focused largely on reducing the transmission of such diseases and on breeding resistant varieties of crop plants.

. Watson was familiar with the type of X-ray diffraction pattern that helical molecules produce, and an examination of the photo that Wilkins showed him confirmed that DNA was helical in shape.

The photo also augmented earlier data obtained by Franklin and others suggesting the width of the helix and the spacing of the nitrogenous bases along it. The pattern in this photo implied that the helix was made up of two strands, contrary to a three-stranded model that Linus Pauling had proposed a short time earlier. The presence of two strands accounts for the now-familiar term double helix. DNA is shown in some of its many different representations in Figure 16.7.

A 50% frequency of recombination in such testcrosses is observed for any two genes that are located on different chromosomes and thus cannot be linked.

The physical basis of recombination between unlinked genes is the random orientation of homologous chromosomes at metaphase I of meiosis, which leads to the independent assortment of the two unlinked genes (see Figure 13.11 and the question in the Figure 15.2 legend).

When 50% of all offspring are recombinants, as in this example, geneticists say that there is a 50% frequency of recombination.

The predicted phenotypic ratios among the offspring are similar to what Mendel actually found in his YyRr * yyrr crosses.

With a normal coin, the chance of tossing heads is 1 ⁄ 2, and the chance of tossing tails is 1 ⁄ 2. The probability of drawing the ace of spades from a 52-card deck is 1 ⁄ 52.

The probabilities of all possible outcomes for an event must add up to 1. With a deck of cards, the chance of picking a card other than the ace of spades is 51⁄ 52.

Completing and Targeting the Functional Protein:

The process of translation is often not sufficient to make a functional protein. In this section, you will learn about modifications that polypeptide chains undergo after the translation process as well as some of the mechanisms used to target completed proteins to specific sites in the cell.

In addition to genes whose products normally promote cell division, cells contain genes whose normal products inhibit cell division. Such genes are called tumor-suppressor genes since the proteins they encode help prevent uncontrolled cell growth. Any mutation that decreases the normal activity of a tumor-suppressor protein may contribute to the onset of cancer, in effect stimulating growth through the absence of suppression.

The protein products of tumor-suppressor genes have various functions. Some repair damaged DNA, a function that prevents the cell from accumulating cancer-causing mutations. Other tumor-suppressor proteins control the adhesion of cells to each other or to the extracellular matrix; proper cell anchorage is crucial in normal tissues—and is often absent in cancers. Still other tumor-suppressor proteins are components of cellsignaling pathways that inhibit the cell cycle.

The purple-flower allele and the white-flower allele are two DNA sequence variations possible at the flower-color locus on a pea plant's chromosomes.

The purple-flower allele sequence allows synthesis of purple pigment, and the white-flower allele sequence does not.

Our ability to extract the intended message from a written language depends on reading the symbols in the correct groupings—that is, in the correct reading frame. Consider this statement: "The red dog ate the bug." Group the letters incorrectly by starting at the wrong point, and the result will probably be gibberish: for example, "her edd oga tet heb ug."

The reading frame is also important in the molecular language of cells. The short stretch of polypeptide shown in Figure 17.5, for instance, will be made correctly only if the mRNA nucleotides are read from left to right (5' -> 3') in the groups of three shown in the figure: UGG UUU GGC UCA. Although a genetic message is written with no spaces between the codons, the cell's proteinsynthesizing machinery reads the message as a series of nonoverlapping three-letter words. The message is not read as a series of overlapping words—UGGUUU, and so on— which would convey a very different message.

Many proteins work together in DNA replication and repair:

The relationship between structure and function is manifest in the double helix. The idea that there is specific pairing of nitrogenous bases in DNA was the flash of inspiration that led Watson and Crick to the double helix. At the same time, they saw the functional significance of the base-pairing rules. They ended their classic paper with this wry statement: "It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material."* In this section, you will learn about the basic principle of DNA replication, as well as some important details of the process.

Termination of Translation: The final stage of translation is termination (Figure 17.21). Elongation continues until a stop codon in the mRNA reaches the A site. The nucleotide base triplets UAG, UAA, and UGA (all written 5¿ S 3¿) do not code for amino acids but instead act as signals to stop translation.A release factor, a protein shaped like an aminoacyl tRNA, binds directly to the stop codon in the A site.

The release factor causes the addition of a water molecule instead of an amino acid to the polypeptide chain. (Water molecules are abundant in the cytosol.) This reaction breaks (hydrolyzes) the bond between the completed polypeptide and the tRNA in the P site, releasing the polypeptide through the exit tunnel of the ribosome's large subunit. The remainder of the translation assembly then comes apart in a multistep process, aided by other protein factors. Breakdown of the translation assembly requires the hydrolysis of two more GTP molecules.

Cystic Fibrosis: The most common lethal genetic disease in the United States is cystic fibrosis, which strikes one out of every 2,500 people of European descent but is much rarer in other groups. Among people of European descent, one out of 25 (4%) are carriers of the cystic fibrosis allele. The normal allele for this gene codes for a membrane protein that functions in the transport of chloride ions between certain cells and the extracellular fluid. These chloride transport channels are defective or absent in the plasma membranes of children who inherit two recessive alleles for cystic fibrosis.

The result is an abnormally high concentration of intracellular chloride, which causes an uptake of water due to osmosis. This in turn causes the mucus that coats certain cells to become thicker and stickier than normal. The mucus builds up in the pancreas, lungs, digestive tract, and other organs, leading to multiple (pleiotropic) effects, including poor absorption of nutrients from the intestines, chronic bronchitis, and recurrent bacterial infections.

Careful examination of a micrograph of the 46 human chromosomes from a single cell in mitosis reveals that there are two chromosomes of each of 23 types. This becomes clear when images of the chromosomes are arranged in pairs, starting with the longest chromosomes.

The resulting ordered display is called a karyotype (Figure 13.3)

Viral Envelopes: An animal virus equipped with an envelope—that is, a membranous outer layer—uses it to enter the host cell. Protruding from the outer surface of this envelope are viral glycoproteins that bind to specific receptor molecules on the surface of a host cell. Figure 19.8 outlines the events in the replicative cycle of an enveloped virus with an RNA genome. Ribosomes bound to the endoplasmic reticulum (ER) of the host cell make the protein parts of the envelope glycoproteins; cellular enzymes in the ER and Golgi apparatus then add the sugars.

The resulting viral glycoproteins, embedded in membrane derived from the host cell, are transported to the cell surface. In a process much like exocytosis, new viral capsids are wrapped in membrane as they bud from the cell. In other words, the viral envelope is usually derived from the host cell's plasma membrane, although all or most of the molecules of this membrane are specified by viral genes. The enveloped viruses are now free to infect other cells. This replicative cycle does not necessarily kill the host cell, in contrast to the lytic cycles of phages

.Mendel tested his seven pea characters in various dihybrid combinations and always observed a 9:3:3:1 phenotypic ratio in the F2 generation. Is this consistent with the 3:1 phenotypic ratio seen for the monohybrid cross shown in Figure 14.5? To explore this question, count the number of yellow and green peas, ignoring shape, and calculate the ratio

The results of Mendel's dihybrid experiments are the basis for what we now call the law of independent assortment, which states that two or more genes assort independently—that is, each pair of alleles segregates independently of any other pair of alleles—during gamete formation.

Translation is the synthesis of a polypeptide using the information in the mRNA. During this stage, there is a change in language: The cell must translate the nucleotide sequence of an mRNA molecule into the amino acid sequence of a polypeptide.

The sites of translation are ribosomes, molecular complexes that facilitate the orderly linking of amino acids into polypeptide chains.

A tRNA molecule consists of a single RNA strand that is only about 80 nucleotides long (compared to hundreds of nucleotides for most mRNA molecules). Because of the presence of complementary stretches of nucleotide bases that can hydrogen-bond to each other, this single strand can fold back on itself and form a molecule with a threedimensional structure. Flattened into one plane to clarify this base pairing, a tRNA molecule looks like a cloverleaf (Figure 17.16a).

The tRNA actually twists and folds into a compact three-dimensional structure that is roughly L-shaped (Figure 17.16b), with the 5¿ and 3¿ ends of the linear tRNA both located near one end of the structure. The protruding 3¿ end acts as the attachment site for an amino acid. The loop extending from the other end of the L includes the anticodon, the particular nucleotide triplet that base-pairs to a specific mRNA codon. Thus, the structure of a tRNA molecule fits its function.

Repressible and Inducible Operons: Two Types of Negative Gene Regulation

The trp operon is said to be a repressible operon because its transcription is usually on but can be inhibited (repressed) when a specific small molecule (in this case, tryptophan) binds allosterically to a regulatory protein. In contrast, an inducible operon is usually off but can be stimulated (induced) to be on when a specific small molecule interacts with a different regulatory protein. The classic example of an inducible operon is the lac operon (lac stands for "lactose").

Differential Gene Expression: A typical human cell might express about 20% of its proteincoding genes at any given time. Highly differentiated cells, such as muscle or nerve cells, express an even smaller fraction of their genes. Almost all the cells in a multicellular organism contain an identical genome. (Cells of the immune system are one exception, as you will see in Chapter 43.) A subset of genes is expressed in each cell type; some of these are "housekeeping" genes, expressed by many cell types, while others are unique to that cell type.

The uniquely expressed genes allow these cells to carry out their specific function. The differences between cell types, therefore, are due not to different genes being present, but to differential gene expression, the expression of different genes by cells with the same genome.

The Evolutionary Significance of Small ncRNAs: Small ncRNAs can regulate gene expression at multiple steps and in many ways. While this section has focused on ncRNAs in eukaryotes, small ncRNAs are also used by bacteria as a defense system, called the CRISPR-Cas9 system, against viruses that infect them. (You'll learn more about this in Concept 19.2.)

The use of ncRNAs thus evolved long ago, but we don't yet know how bacterial ncRNAs are related to those of eukaryotes

The immune system is a critical part of the body's natural defenses (see Chapter 43). It is also the basis for the major medical tool used to prevent viral infections—vaccines. A vaccine is a harmless derivative of a pathogen that stimulates the immune system to mount defenses against the harmful pathogen. Smallpox, a viral disease that was once a devastating scourge in many parts of the world, was eradicated by a vaccination program carried out by the World Health Organization (WHO).

The very narrow host range of the smallpox virus—it infects only humans—was a critical factor in the success of this program. Similar worldwide vaccination campaigns are currently under way to eradicate polio and measles. Effective vaccines are also available to protect against rubella, mumps, hepatitis B, and a number of other viral diseases.

Genomic imprinting may affect only a small fraction of the genes in mammalian genomes, but most of the known imprinted genes are critical for embryonic development. In experiments with mice, embryos engineered to inherit both copies of certain chromosomes from the same parent usually die before birth, whether that parent is male or female. A few years ago, however, scientists in Japan combined the genetic material from two eggs in a zygote while allowing expression of the Igf2 gene, along with several other imprinted genes, from only one of the egg nuclei.

The zygote developed into an apparently healthy mouse. Normal development seems to require that embryonic cells have exactly one active copy—not zero, not two—of certain genes. The association of improper imprinting with abnormal development and certain cancers has stimulated ongoing studies of how different genes are imprinted.

(The regulatory role of DNA methylation is discussed in Concept 18.2.) A particular region of each X chromosome contains several genes involved in the inactivation process. The two regions, one on each X chromosome, associate briefly with each other in each cell at an early stage of embryonic development.

Then one of the genes, called XIST (for X-inactive specific transcript), becomes active only on the chromosome that will become the Barr body. Multiple copies of the RNA product of this gene apparently attach to the X chromosome on which they are made, eventually almost covering it. Interaction of this RNA with the chromosome initiates X inactivation, and the RNA products of nearby genes help to regulate the process.

The Drosophila egg develops in one of the female's ovaries, adjacent to ovarian cells called nurse cells and surrounded by so-called follicle cells (Figure 18.19b, top). These support cells supply the egg with nutrients, mRNAs, and other substances needed for development and make the egg shell. After fertilization and laying of the egg, embryonic development results in the formation of a segmented larva, which goes through three larval stages.

Then, in a process much like that by which a caterpillar becomes a butterfly, the fly larva forms a pupa in which it metamorphoses into the adult fly pictured in Figure 18.19a.

Since 1993, a number of research studies have uncovered microRNAs (miRNAs)—small, single-stranded RNA molecules capable of binding to complementary sequences in mRNA molecules. A longer RNA precursor is processed by cellular enzymes into an miRNA, a single-stranded RNA of about 22 nucleotides that forms a complex with one or more proteins (Figure 18.14). The miRNA allows the complex to bind to any mRNA molecule with at least 7 or 8 nucleotides of complementary sequence. The miRNA-protein complex then degrades the target mRNA or, less often, simply blocks its translation.

There are approximately 1,500 genes for miRNAs in the human genome, and biologists estimate that expression of at least one-half of all human genes may be regulated by miRNAs, a remarkable figure given that the existence of miRNAs was unknown 25 years ago.

Tests for Identifying Carriers: Most children with recessive disorders are born to parents with normal phenotypes. The key to accurately assessing the genetic risk for a particular disease is therefore to find out whether the prospective parents are heterozygous carriers of the recessive allele. For an increasing number of heritable disorders, tests are available that can distinguish individuals of normal phenotype who are dominant homozygotes from those who are heterozygous carriers.

There are now tests that can identify carriers of the alleles for Tay-Sachs disease, sickle-cell disease, and the most common form of cystic fibrosis. A program testing for carriers of Tay-Sachs disease that began in the 1980s has successfully reduced the rate of babies born with this disease.

Another advantage of the fruit fly is that it has only four pairs of chromosomes, which are easily distinguishable with a light microscope.

There are three pairs of autosomes and one pair of sex chromosomes. Female fruit flies have a pair of homologous X chromosomes, and males have one X chromosome and one Y chromosome.

Viral Genetic Material: Table 19.1 shows the common classification system for animal viruses, which is based on their genetic material: double- or single-stranded DNA, or double- or single-stranded RNA. Although some phages and most plant viruses are RNA viruses, the broadest variety of RNA genomes is found among the viruses that infect animals.

There are three types of singlestranded RNA genomes found in animal viruses (classes IV-VI in Table 19.1). The genome of class IV viruses can directly serve as mRNA and thus can be translated into viral protein immediately after infection. Figure 19.8 shows a virus of class V, in which the RNA genome serves instead as a template for mRNA synthesis. The RNA genome is transcribed into complementary RNA strands, which function both as mRNA and as templates for the synthesis of additional copies of genomic RNA. All viruses that use an RNA genome as a template for mRNA transcription require RNA S RNA synthesis. These viruses use a viral enzyme capable of carrying out this process; there are no such enzymes in most cells. The enzyme used in this process is encoded by the viral genome, and after its synthesis the protein is packaged during viral self-assembly with the genome inside the viral capsid.

The Roles of General and Specific Transcription Factors:

There are two types of transcription factors: General transcription factors act at the promoter of all genes, while some genes require specific transcription factors that bind to control elements that may be close to or farther away from the promoter.

When we use Mendel's laws to predict possible outcomes of matings, it is important to remember that each child represents an independent event in the sense that its genotype is unaffected by the genotypes of older siblings. Suppose that John and Carol have three more children, and all three have the hypothetical hereditary disease.

There is only one chance in 64 (1 ⁄ 4 * 1 ⁄ 4 * 1 ⁄ 4) that such an outcome will occur. Despite this run of misfortune, the chance that a fourth child of this couple will have the disease remains 1 ⁄ 4.

Some viruses have envelopes that are not derived from plasma membrane. Herpesviruses, for example, are temporarily cloaked in membrane derived from the nuclear envelope of the host; they then shed this membrane in the cytoplasm and acquire a new envelope made from membrane of the Golgi apparatus. These viruses have a double-stranded DNA genome and replicate within the host cell nucleus, using a combination of viral and cellular enzymes to replicate and transcribe their DNA. In the case of herpesviruses, copies of the viral DNA can remain behind as mini-chromosomes in the nuclei of certain nerve cells.

There they remain latent until some sort of physical or emotional stress triggers a new round of active virus production. The infection of other cells by these new viruses causes the blisters characteristic of herpes, such as cold sores or genital sores. Once someone acquires a herpesvirus infection, flare-ups may recur throughout the person's life.

Once a SNP is identified that is found in all affected people, researchers focus on that region and sequence it. In nearly all cases, the SNP itself does not contribute directly to the disease in question by altering the encoded protein; in fact, most SNPs are in noncoding regions. Instead, if the SNP and a disease-associated allele are close enough to be genetically linked, scientists can take advantage of the fact that crossing over between the marker and the gene is very unlikely during gamete formation.

Therefore, the marker and gene will almost always be inherited together, even though the marker is not part of the gene (Figure 20.15). SNPs have been found that correlate with diabetes, heart disease, and several types of cancer, and the search is on for genes that might be involved.

We have noted that merely saying a gene codes for a polypeptide is an oversimplification. Most eukaryotic genes contain noncoding segments (such as introns), so large portions of these genes have no corresponding segments in polypeptides. Molecular biologists also often include promoters and certain other regulatory regions of DNA within the boundaries of a gene.

These DNA sequences are not transcribed, but they can be considered part of the functional gene because they must be present for transcription to occur. Our definition of a gene must also be broad enough to include the DNA that is transcribed into rRNA, tRNA, and other RNAs that are not translated. These genes have no polypeptide products but play crucial roles in the cell. Thus, we arrive at the following definition: A gene is a region of DNA that can be expressed to produce a final functional product that is either a polypeptide or an RNA molecule.

When a phage infects a bacterial cell that has the CRISPRCas system, the DNA of the invading phage is integrated into the genome between two repeat sequences. If the cell survives the infection, any further attempt by the same type of phage to infect this cell (or its offspring) triggers transcription of the CRISPR region into RNA molecules (Figure 19.7).

These RNAs are cut into pieces and then bound by Cas proteins. The Cas protein uses a portion of the phage-related RNA as a homing device to identify the invading phage DNA and cut it, leading to its destruction. In Concept 20.1, you'll learn how this system is used in the laboratory to alter genes in other cells.

Polygenic Inheritance: Mendel studied characters that could be classified on an either-or basis, such as purple versus white flower color. But many characters, such as human skin color and height, are not one of two discrete characters, but instead vary in the population in gradations along a continuum.

These are called quantitative characters. Quantitative variation usually indicates polygenic inheritance, an additive effect of two or more genes on a single phenotypic character. (In a way, this is the converse of pleiotropy, where a single gene affects several phenotypic characters.)

Prions: Proteins as Infectious Agents The viruses we have discussed in this chapter are infectious agents that spread diseases, and their genetic material is composed of nucleic acids, whose ability to be replicated is well known. Surprisingly, there are also proteins that are known to be infectious. Proteins called prions appear to cause a number of degenerative brain diseases in various animal species.

These diseases include scrapie in sheep; mad cow disease, which has plagued the European beef industry in recent years; and Creutzfeldt-Jakob disease in humans, which has caused the death of some 175 people in the United Kingdom since 1996. Prions can be transmitted in food, as may occur when people eat prion-laden beef from cattle with mad cow disease. Kuru, another human disease caused by prions, was identified in the early 1900s among the South Fore natives of New Guinea. A kuru epidemic peaked there in the 1960s, puzzling scientists, who at first thought the disease had a genetic basis.

Multiple Alleles: Only two alleles exist for the pea characters that Mendel studied, but most genes exist in more than two allelic forms. The ABO blood groups in humans, for instance, are determined by that person's two alleles of the blood group gene; there are three possible alleles: I A, IB , and i. A person's blood group may be one of four types: A, B, AB, or O.

These letters refer to two carbohydrates—A and B—that may be found attached to specific cell-surface molecules on red blood cells. An individual's blood cells may have carbohydrate A (type A blood), carbohydrate B (type B), both (type AB), or neither (type O), as shown in Figure 14.11. Matching compatible blood groups is critical for safe blood transfusions (see Concept 43.3).

Mechanisms of Post-Transcriptional Regulation: Transcription alone does not constitute gene expression. The expression of a protein-coding gene is ultimately measured by the amount of functional protein a cell makes, and much happens between the synthesis of the RNA transcript and the activity of the protein in the cell. Many regulatory mechanisms operate at the various stages after transcription (see Figure 18.6).

These mechanisms allow a cell to rapidly finetune gene expression in response to environmental changes without altering its transcription patterns. Here we discuss how cells can regulate gene expression once a gene has been transcribed

How can binding of activators to an enhancer located far from the promoter influence transcription? One study shows that the proteins regulating a mouse globin gene contact both the gene's promoter and an enhancer located about 50,000 nucleotides upstream. This and many other studies support the currently accepted model, in which proteinmediated bending of the DNA is thought to bring the bound activators into contact with a group of mediator proteins, which in turn interact with general transcription factors at the promoter (Figure 18.10).

These protein-protein interactions help assemble and position the initiation complex on the promoter, and allow the promoter and enhancer to come together in a very specific fashion, in spite of what is often a large number of nucleotide pairs between them. In the Scientific Skills Exercise, you can work with data from an experiment that identified the control elements in an enhancer of a particular human gene.

To elongate the other new strand of DNA in the mandatory 5′ S 3′ direction, DNA pol III must work along the other template strand in the direction away from the replication fork. The DNA strand elongating in this direction is called the lagging strand. In contrast to the leading strand, which elongates continuously, the lagging strand is synthesized discontinuously, as a series of segments.

These segments of the lagging strand are called Okazaki fragments, after Reiji Okazaki, the Japanese scientist who discovered them. The fragments are about 1,000-2,000 nucleotides long in E. coli and 100-200 nucleotides long in eukaryotes. Figure 16.16 illustrates the steps in the synthesis of the lagging strand at one fork. Whereas only one primer is required on the leading strand, each Okazaki fragment on the lagging strand must be primed separately (steps 1 and 4 ). After DNA pol III forms an Okazaki fragment (steps 2 to 4 ), another DNA polymerase, DNA pol I, replaces the RNA nucleotides of the adjacent primer with DNA nucleotides one at a time (step 5 ). But DNA pol I cannot join the final nucleotide of this replacement DNA segment to the first DNA nucleotide of the adjacent Okazaki fragment. Another enzyme, DNA ligase, accomplishes this task, joining the sugar-phosphate backbones of all the Okazaki fragments into a continuous DNA strand (step 6 ).

Histone Modifications and DNA Methylation: There is abundant evidence that chemical modifications to histones, found in all eukaryotic organisms, play a direct role in the regulation of gene transcription. The N-terminus of each histone protein in a nucleosome protrudes outward from the nucleosome (Figure 18.7a).

These so-called histone tails are accessible to various modifying enzymes that catalyze the addition or removal of specific chemical groups, such as acetyl (¬COCH3), methyl, and phosphate groups (see Figure 4.9). Generally, histone acetylation—the addition of an acetyl group to an amino acid in a histone tail—appears to promote transcription by opening up the chromatin structure (Figure 18.7b), while the addition of methyl groups to histones can lead to the condensation of chromatin and reduced transcription. Often, the addition of a particular chemical group may create a new binding site for enzymes that further modify chromatin structure in various ways.

Nuclear Architecture and Gene Expression: You saw in Figure 16.23b that each chromosome in the interphase nucleus occupies a distinct territory. The chromosomes are not completely isolated, however. Recently, chromosome conformation capture (3C) techniques have been developed that allow researchers to cross-link and identify regions of chromosomes that associate with each other during interphase.

These studies reveal that loops of chromatin extend from individual chromosomal territories into specific sites in the nucleus (Figure 18.12). Different loops from the same chromosome and loops from other chromosomes may congregate in such sites, some of which are rich in RNA polymerases and other transcription-associated proteins. Like a recreation center that draws members from many different neighborhoods, these so-called transcription factories are thought to be areas specialized for a common function.

Specific sequences of nucleotides along the DNA mark where transcription of a gene begins and ends. The DNA sequence where RNA polymerase attaches and initiates transcription is known as the promoter; in bacteria, the sequence that signals the end of transcription is called the terminator. (The termination mechanism is different in eukaryotes; we'll describe it later.) Molecular biologists refer to the direction of transcription as "downstream" and the other direction as "upstream.

These terms are also used to describe the positions of nucleotide sequences within the DNA or RNA. Thus, the promoter sequence in DNA is said to be upstream from the terminator. The stretch of DNA downstream from the promoter that is transcribed into an RNA molecule is called a transcription unit

From these results, Gurdon concluded that something in the nucleus does change as animal cells differentiate. In frogs and most other animals, nuclear potential tends to be restricted more and more as embryonic development and cell differentiation progress.

These were foundational experiments that ultimately led to stem cell technology, and Gurdon received the 2012 Nobel Prize in Medicine for this work.

In 1843, at the age of 21, Mendel entered an Augustinian monastery, a reasonable choice at that time for someone who valued the life of the mind. He considered becoming a teacher but failed the necessary examination. In 1851, he left the monastery to pursue two years of study in physics and chemistry at the University of Vienna.

These were very important years for Mendel's development as a scientist, in large part due to the strong influence of two professors. One was the physicist Christian Doppler, who encouraged his students to learn science through experimentation and trained Mendel to use mathematics to help explain natural phenomena. The other was a botanist named Franz Unger, who aroused Mendel's interest in the causes of variation in plants.

Some viruses have accessory structures that help them infect their hosts. For instance, a membranous envelope surrounds the capsids of influenza viruses and many other viruses found in animals (Figure 19.3c). These viral envelopes, which are derived from the membranes of the host cell, contain host cell phospholipids and membrane proteins.

They also contain proteins and glycoproteins of viral origin. (Glycoproteins are proteins with carbohydrates covalently attached.) Some viruses carry a few viral enzyme molecules within their capsids. Many of the most complex capsids are found among the viruses that infect bacteria, called bacteriophages, or simply phages. The first phages studied included seven that infect Escherichia coli. These seven phages were named type 1 (T1), type 2 (T2), and so forth, in the order of their discovery. The three "T-even" phages (T2, T4, and T6) turned out to be very similar in structure. Their capsids have elongated icosahedral heads enclosing their DNA. Attached to the head is a protein tail piece with fibers by which the phages attach to a bacterial cell (Figure 19.3d). In the next section, we'll examine how these few viral parts function together with cellular components to produce large numbers of viral progeny

Viruses replicate only in host cells: Viruses lack metabolic enzymes and equipment for making proteins, such as ribosomes.

They are obligate intracellular parasites; in other words, they can replicate only within a host cell. It is fair to say that viruses in isolation are merely packaged sets of genes in transit from one host cell to another.

Nutritional Mutants in Neurospora: Scientific Inquiry A breakthrough came a few years later at Stanford University, where Beadle and Edward Tatum began working with a bread mold, Neurospora crassa, a haploid species. To observe a change in a mutant's phenotype, Beadle and Tatum needed to disable just one allele (rather than two, as in a diploid species) of a protein-coding gene required for a specific metabolic activity.

They bombarded Neurospora with X-rays, known to cause mutations, and looked among the survivors for mutants that differed in their nutritional needs from the wild-type bread mold.

Hershey and Chase found that the phage DNA entered the host cells but the phage protein did not. Moreover, when these bacteria were returned to a culture medium and the infection ran its course, the E. coli released phages that contained some radioactive phosphorus. This result further showed that the DNA inside the cell played an ongoing role during the infection process.

They concluded that the DNA injected by the phage must be the molecule carrying the genetic information that makes the cells produce new viral DNA and proteins. The Hershey-Chase experiment was a landmark study because it provided powerful evidence that nucleic acids, rather than proteins, are the hereditary material, at least for certain viruses.

Three properties of RNA enable some RNA molecules to function as enzymes. First, because RNA is single-stranded, a region of an RNA molecule may base-pair, in an antiparallel arrangement, with a complementary region elsewhere in the same molecule; this gives the molecule a particular threedimensional structure. A specific structure is essential to the catalytic function of ribozymes, just as it is for enzymatic proteins. Second, like certain amino acids in an enzymatic protein, some of the bases in RNA contain functional groups that can participate in catalysis.

Third, the ability of RNA to hydrogenbond with other nucleic acid molecules (either RNA or DNA) adds specificity to its catalytic activity. For example, complementary base pairing between the RNA of the spliceosome and the RNA of a primary RNA transcript precisely locates the region where the ribozyme catalyzes splicing. Later in this chapter, you will see how these properties of RNA also allow it to perform important noncatalytic roles in the cell, such as recognition of the three-nucleotide codons on mRNA.

Researchers have developed a variety of methods to test the mutagenic activity of chemicals. A major application of these tests is the preliminary screening of chemicals to identify those that may cause cancer.

This approach makes sense because most carcinogens (cancer-causing chemicals) are mutagenic, and conversely, most mutagens are carcinogenic.

How does the E. coli cell sense the glucose concentration and relay this information to the lac operon? Again, the mechanism depends on the interaction of an allosteric regulatory protein with a small organic molecule, cyclic AMP (cAMP) in this case, which accumulates when glucose is scarce (see Figure 11.11 for the structure of cAMP). The regulatory protein, called cAMP receptor protein (CRP), is an activator, a protein that binds to DNA and stimulates transcription of a gene. When cAMP binds to this regulatory protein, CRP assumes its active shape and can attach to a specific site at the upstream end of the lac promoter (Figure 18.5a).

This attachment increases the affinity of RNA polymerase for the lac promoter, which is actually rather low even when no lac repressor is bound to the operator. By facilitating the binding of RNA polymerase to the promoter and thereby increasing the rate of transcription of the lac operon, the attachment of CRP to the promoter directly stimulates gene expression. Therefore, this mechanism qualifies as positive regulation.

Even with only a dozen control element sequences available, a very large number of combinations is possible. Each combination of control elements is able to activate transcription only when the appropriate activator proteins are present, which may occur at a precise time during development or in a particular cell type. Figure 18.11 illustrates how the use of different combinations of just a few control elements can allow differential regulation of transcription in two representative cell types—liver cells and lens cells.

This can occur because each cell type contains a different group of activator proteins. How cell types come to differ during this process, even though they all arise from one cell (the fertilized egg), will be explored in Concept 18.4.

To summarize: Genes program protein synthesis via genetic messages in the form of messenger RNA. Put another way, cells are governed by a molecular chain of command with a directional flow of genetic information:

This concept was dubbed the central dogma by Francis Crick in 1956. But in the 1970s, scientists were surprised to discover some enzymes that use RNA molecules as templates for DNA synthesis (which we'll cover in Concept 19.2). However, these exceptions do not invalidate the idea that, in general, genetic information flows from DNA to RNA to protein. In the next section, we discuss how the instructions for assembling amino acids into a specific order are encoded in nucleic acids.

It is clear that alternative RNA splicing can significantly expand the repertoire of a eukaryotic genome. In fact, alternative splicing was proposed as one explanation for the surprisingly low number of human genes counted when the human genome was sequenced. The number of human genes was found to be similar to that of a soil worm (nematode), a mustard plant, or a sea anemone.

This discovery prompted questions about what, if not the number of genes, accounts for the more complex morphology (external form) of humans. It turns out that more than 90% of human protein-coding genes probably undergo alternative splicing. Thus, the extent of alternative splicing greatly multiplies the number of possible human proteins, which may be better correlated with complexity of form.

The Discovery of Viruses: Scientific Inquiry Tobacco mosaic disease stunts the growth of tobacco plants and gives their leaves a mottled, or mosaic, coloration. In 1883, Adolf Mayer, a German scientist, discovered that he could transmit the disease from plant to plant by rubbing sap extracted from diseased leaves onto healthy plants. After an unsuccessful search for an infectious microbe in the sap, Mayer suggested that the disease was caused by unusually small bacteria that were invisible under a microscope.

This hypothesis was tested a decade later by Dmitri Ivanowsky, a Russian biologist who passed sap from infected tobacco leaves through a filter designed to remove bacteria. After filtration, the sap still produced mosaic disease.

At one time, the onset of symptoms was the only way to know if a person had inherited the Huntington's allele, but this is no longer the case. By analyzing DNA samples from a large family with a high incidence of the disorder, geneticists tracked the Huntington's allele to a locus near the tip of chromosome 4, and the gene was sequenced in 1993.

This information led to the development of a test that could detect the presence of the Huntington's allele in an individual's genome. (The methods that make such tests possible are discussed in Concepts 20.1 and 20.4.) The availability of this test poses an agonizing dilemma for those with a family history of Huntington's disease. Some individuals may want to be tested for this disease, whereas others may decide it would be too stressful to find out.

General Transcription Factors at the Promoter:

To initiate transcription, eukaryotic RNA polymerase requires the assistance of transcription factors. Some transcription factors (such as those illustrated in Figure 17.9) are essential for the transcription of all protein-coding genes; therefore, they are often called general transcription factors. A few general transcription factors bind to a DNA sequence such as the TATA box within the promoter, but many bind to proteins, including other transcription factors and RNA polymerase II.

In the last 15 years, "next-generation sequencing" techniques have been developed that are much faster (Figure 20.2). DNA fragments are amplified (copied) to yield an enormous number of identical fragments (Figure 20.3). A specific strand of each fragment is immobilized, and the complementary strand is synthesized, one nucleotide at a time. A chemical technique enables electronic monitors to identify in real time which of the four nucleotides is added; this method is thus called sequencing by synthesis. Thousands or hundreds of thousands of fragments,each about 300 nucleotides long, are sequenced in parallel in machines like those shown in Figure 20.2, accounting for the high rate of nucleotides sequenced per hour.

This is an example of "high-throughput" DNA technology and is currently the method of choice for studies where massive numbers of DNA samples—even a set of numerous fragments representing an entire genome—are being sequenced.

Proofreading and Repairing DNA: We cannot attribute the accuracy of DNA replication solely to the specificity of base pairing. Initial pairing errors between incoming nucleotides and those in the template strand occur at a rate of one in 105 nucleotides. However, errors in the completed DNA molecule amount to only one in 1010 (10 billion) nucleotides, an error rate that is 100,000 times lower.

This is because during DNA replication, many DNA polymerases proofread each nucleotide against its template as soon as it is covalently bonded to the growing strand. Upon finding an incorrectly paired nucleotide, the polymerase removes the nucleotide and then resumes synthesis. (This action is similar to fixing a texting error by deleting the wrong letter and then entering the correct one.)

When a disease-causing recessive allele is rare, it is relatively unlikely that two carriers of the same harmful allele will meet and mate. The probability of passing on recessive traits increases greatly, however, if the man and woman are close relatives (for example, siblings or first cousins).

This is because people with recent common ancestors are more likely to carry the same recessive alleles than are unrelated people.

Termination of Transcription: Bacteria and eukaryotes differ in the way they terminate transcription. In bacteria, transcription proceeds through a terminator sequence in the DNA. The transcribed terminator (an RNA sequence) functions as the termination signal, causing the polymerase to detach from the DNA and release the transcript, which requires no further modification before translation. In eukaryotes, RNA polymerase II transcribes a sequence on the DNA called the polyadenylation signal sequence, which specifies a polyadenylation signal (AAUAAA) in the pre-mRNA.

This is called a "signal" because once this stretch of six RNA nucleotides appears, it is immediately bound by certain proteins in the nucleus. Then, at a point about 10-35 nucleotides downstream from the AAUAAA, these proteins cut the RNA transcript free from the polymerase, releasing the pre-mRNA. The pre-mRNA then undergoes processing, the topic of the next section. Although that cleavage marks the end of the mRNA, the RNA polymerase II continues to transcribe. Enzymes begin to degrade the RNA starting at its newly exposed 5¿ end. The polymerase continues transcribing, pursued by the enzymes, until they catch up to the polymerase and it dissociates from the DNA.

In another application of CRISPR-Cas9, scientists are attempting to address the global problem of insect-borne diseases by altering genes in the insect so that, for example, it cannot transmit disease. An extra twist to this approach is engineering the new allele so that it is much more highly favored for inheritance than is the wild-type allele.

This is called a gene drive because the biased inheritance of the engineered gene during reproduction rapidly "drives" the new allele through the population.

Because each pair of homologous chromosomes is positioned independently of the other pairs at metaphase I, the first meiotic division results in each pair sorting its maternal and paternal homologs into daughter cells independently of every other pair.

This is called independent assortment. Each daughter cell represents one outcome of all possible combinations of maternal and paternal chromosomes.

. Suppose that the couple represented in the second generation of Figure 14.15 decides to have one more child. What is the probability that the child will have a widow's peak?

This is equivalent to a Mendelian F1 monohybrid cross (Ww * Ww), and therefore the probability that a child will inherit a dominant allele and have a widow's peak is 3 ⁄ 4 (1 ⁄ 4 WW + 1 ⁄ 2 Ww).

In 2015, after a year of use and review by scientists, the first nanopore sequencer went on the market; this device is the size of a small candy bar and connects to a computer via a USB port. Associated software allows immediate identification and analysis of the sequence.

This is only one of many approaches to further increase the rate and cut the cost of sequencing, while also allowing the methodology to move out of the laboratory and into the field.

In a typical breeding experiment, Mendel cross-pollinated two contrasting, true-breeding pea varieties—for example, purple-flowered plants and white-flowered plants (see Figure 14.2).

This mating, or crossing, of two true-breeding varieties is called hybridization. The true-breeding parents are referred to as the P generation (parental generation), and their hybrid offspring are the F1 generation (first filial generation, the word filial from the Latin word for "son"). Allowing these F1 hybrids to self-pollinate (or to cross-pollinate with other F1 hybrids) produces an F2 generation (second filial generation). Mendel usually followed traits for at least the P, F1, and F2 generations.

The term lysogenic signifies that prophages are capable of generating active phages that lyse their host cells.

This occurs when the l genome (or that of another temperate phage) is induced to exit the bacterial chromosome and initiate a lytic cycle. An environmental signal, such as a certain chemical or high-energy radiation, usually triggers the switchover from the lysogenic to the lytic mode.

The polypeptides of proteins destined for the endomembrane system or for secretion are marked by a signal peptide, which targets the protein to the ER (Figure 17.22). The signal peptide, a sequence of about 20 amino acids at or near the leading end (N-terminus) of the polypeptide, is recognized as it emerges from the ribosome by a protein-RNA complex called a signal-recognition particle (SRP).

This particle escorts the ribosome to a receptor protein built into the ER membrane. The receptor is part of a multiprotein translocation complex. Polypeptide synthesis continues there, and the growing polypeptide snakes across the membrane into the ER lumen via a protein pore. The rest of the completed polypeptide, if it is to be secreted from the cell, is released into solution within the ER lumen. Alternatively, if the polypeptide is to be a membrane protein, it remains partially embedded in the ER membrane. In either case, it travels in a transport vesicle to its destination (see, for example, Figure 7.9).

Types of influenza often emerge as outbreaks of illness. In 2009, a widespread outbreak, or epidemic, of a flu-like illness appeared in Mexico and the United States. The infectious agent was quickly identified as an influenza virus related to viruses that cause the seasonal flu.

This particular virus was named H1N1 for reasons that will be explained shortly. The illness spread rapidly, prompting WHO to declare a global epidemic, or pandemic, shortly thereafter. Half a year later, the disease had reached 207 countries, infecting over 600,000 people and killing almost 8,000.

Bicoid: A Morphogen That Determines Head Structures To see how maternal effect genes determine the body axes of the offspring, we will focus on one such gene, called bicoid, a term meaning "two-tailed." An embryo or larva whose mother has two mutant bicoid alleles lacks the front half of its body and has posterior structures at both ends (Figure 18.21).

This phenotype suggested to Nüsslein-Volhard and her colleagues that the product of the mother's bicoid gene is essential for setting up the anterior end of the fly and might be concentrated at the future anterior end of the embryo. This hypothesis is an example of the morphogen gradient hypothesis first proposed by embryologists a century ago, in which gradients of substances called morphogens establish an embryo's axes and other features of its form

In contrast, the eukaryotic cell's nuclear envelope segregates transcription from translation and provides a compartment for extensive RNA processing.

This processing stage includes additional steps, discussed earlier, the regulation of which can help coordinate the eukaryotic cell's elaborate activities. Figure 17.25 summarizes the path from gene to polypeptide in a eukaryotic cell.

DNA Replication: A Closer Look The bacterium E. coli has a single chromosome of about 4.6 million nucleotide pairs. In a favorable environment, an E. coli cell can copy all of this DNA and divide to form two genetically identical daughter cells in considerably less than an hour. Each of your somatic cells has 46 DNA molecules in its nucleus, one long double-helical molecule per chromosome. In all, that represents about 6 billion nucleotide pairs, or over 1,000 times more DNA than is found in most bacterial cells. If we were to print the one-letter symbols for these bases (A, G, C, and T) the size of the type you are now reading, the 6 billion nucleotide pairs of information in a diploid human cell would fill about 1,400 biology textbooks. Yet it takes one of your cells just a few hours to copy all of this DNA during S phase of interphase.

This replication of an enormous amount of genetic information is achieved with very few errors—only about one per 10 billion nucleotides. The copying of DNA is remarkable in its speed and accuracy. More than a dozen enzymes and other proteins participate in DNA replication. Much more is known about how this "replication machine" works in bacteria (such as E. coli) than in eukaryotes, and we will describe the basic steps of the process for E. coli, except where otherwise noted. What scientists have learned about eukaryotic DNA replication suggests, however, that most of the process is fundamentally similar for prokaryotes and eukaryotes.

Biologists are excited about these discoveries, which have revealed a large and diverse population of RNA molecules in the cell that play crucial roles in regulating gene expression— but have gone largely unnoticed until fairly recently. Our long-standing view that because mRNAs code for proteins, they are the most important RNAs functioning in the cell demands revision.

This represents a major shift in the thinking of biologists, one that you are witnessing as students entering this field of study.

This model of DNA replication remained untested for several years following publication of the DNA structure. The requisite experiments were simple in concept but difficult to perform. Watson and Crick's model predicts that when a double helix replicates, each of the two daughter molecules will have one old strand, from the parental molecule, and one newly made strand.

This semiconservative model can be distinguished from a conservative model of replication, in which the two parental strands somehow come back together after the process (that is, the parental molecule is conserved). In yet a third model, called the dispersive model, all four strands of DNA following replication have a mixture of old and new DNA (Figure 16.10).

Among the most useful genetic markers in tracking down genes that contribute to diseases and disorders are single base-pair variations in the genomes of the human population. A single base-pair site where variation is found in at least 1% of the population is called a single nucleotide polymorphism (SNP, pronounced "snip"). A few million SNPs occur in the human genome, about once in 100-300 base pairs of both coding and noncoding DNA sequences.

To find SNPs in large numbers of people, it isn't necessary to sequence their DNA; SNPs can be detected by very sensitive microarray assays, RNA-seq, or PCR.

To clone pieces of DNA using bacteria, researchers first obtain a plasmid (originally isolated from a bacterial cell and genetically engineered for efficient cloning) and insert DNA from another source ("foreign" DNA) into it (Figure 20.4). The resulting plasmid is now a recombinant DNA molecule, a molecule containing DNA from two different sources, very often different species. The plasmid is then returned to a bacterial cell, producing a recombinant bacterium.

This single cell reproduces through repeated cell divisions to form a clone of cells, a population of genetically identical cells. Because the dividing bacteria replicate the recombinant plasmid and pass it on to their descendants, the foreign DNA and any genes it carries are cloned at the same time. The production of multiple copies of a single gene is a type of DNA cloning called gene cloning.

Considering all that can occur in the pathway from genotype to phenotype, it is indeed impressive that Mendel could uncover the fundamental principles governing the transmission of individual genes from parents to offspring. Mendel's laws of segregation and of independent assortment explain heritable variations in terms of alternative forms of genes (hereditary "particles," now known as the alleles of genes) that are passed along, generation after generation, according to simple rules of probability.

This theory of inheritance is equally valid for peas, flies, fishes, birds, and human beings—indeed, for any organism with a sexual life cycle. Furthermore, by extending the principles of segregation and independent assortment to help explain such hereditary patterns as epistasis and quantitative characters, we begin to see how broadly Mendelian genetics applies. From Mendel's abbey garden came a theory of particulate inheritance that anchors modern genetics. In the last section of this chapter, we will apply Mendelian genetics to human inheritance, with emphasis on the transmission of hereditary diseases.

Transcription is the synthesis of RNA using information in the DNA. The two nucleic acids are written in different forms of the same language, and the information is simply transcribed, or "rewritten," from DNA to RNA. Just as a DNA strand provides a template for making a new complementary strand during DNA replication (see Concept 16.2), it also can serve as a template for assembling a complementary sequence of RNA nucleotides. For a protein-coding gene, the resulting RNA molecule is a faithful transcript of the gene's protein building instructions.

This type of RNA molecule is called messenger RNA (mRNA) because it carries a genetic message from the DNA to the protein-synthesizing machinery of the cell. (Transcription is the general term for the synthesis of any kind of RNA on a DNA template. Later, you will learn about some other types of RNA produced by transcription.)

In general, genetic disorders are not evenly distributed among all groups of people. For example, the incidence of Tay-Sachs disease, which we described earlier in this chapter, is disproportionately high among Ashkenazic Jews, Jewish people whose ancestors lived in central Europe. In that population, Tay-Sachs disease occurs in one out of 3,600 births, an incidence about 100 times greater than that among non-Jews or Mediterranean (Sephardic) Jews.

This uneven distribution results from the different genetic histories of the world's peoples during less technological times, when populations were more geographically (and hence genetically) isolated.

Note that either haploid or diploid cells can divide by mitosis, depending on the type of life cycle. Only diploid cells, however, can undergo meiosis because haploid cells have only a single set of chromosomes that cannot be further reduced.

Though the three types of sexual life cycles differ in the timing of meiosis and fertilization, they share a fundamental result: genetic variation among offspring.

:Recessively Inherited Disorders:

Thousands of genetic disorders are known to be inherited as simple recessive traits. These disorders range in severity from relatively mild, such as albinism (lack of pigmentation, which results in susceptibility to skin cancers and vision problems), to life-threatening, such as cystic fibrosis.

Counseling Based on Mendelian Genetics and Probability Rules: Consider the case of a hypothetical couple, John and Carol. Each had a brother who died from the same recessively inherited lethal disease. Before conceiving their first child, John and Carol seek genetic counseling to determine the risk of having a child with the disease. From the information about their brothers, we know that both parents of John and both parents of Carol must have been carriers of the recessive allele.

Thus, John and Carol are both products of Aa * Aa crosses, where a symbolizes the allele that causes this particular disease. We also know that John and Carol are not homozygous recessive (aa), because they do not have the disease. Therefore, their genotypes are either AA or Aa.

Solving Complex Genetics Problems with the Rules of Probability: We can also apply the rules of probability to predict the outcome of crosses involving multiple characters. Recall that each allelic pair segregates independently during gamete formation (the law of independent assortment)

Thus, a dihybrid or other multicharacter cross is equivalent to two or more independent monohybrid crosses occurring simultaneously. By applying what we have learned about monohybrid crosses, we can determine the probability of specific genotypes occurring in the F2 generation without having to construct unwieldy Punnett squares

Second, for each character, an organism inherits two copies (that is, two alleles) of a gene, one from each parent. Remarkably, Mendel made this deduction without knowing about the role, or even the existence, of chromosomes. Each somatic cell in a diploid organism has two sets of chromosomes, one set inherited from each parent (see Figure 13.4).

Thus, a genetic locus is actually represented twice in a diploid cell, once on each homolog of a specific pair of chromosomes. The two alleles at a particular locus may be identical, as in the true-breeding plants of Mendel's P generation. Or the alleles may differ, as in the F1 hybrids (see Figure 14.4).

Extending our analysis to yet another level, we find that heterozygous individuals produce equal numbers of normal and dysfunctional enzyme molecules.

Thus, at the molecular level, the normal allele and the Tay-Sachs allele are codominant. As you can see, whether alleles appear to be completely dominant, incompletely dominant, or codominant depends on the level at which the phenotype is analyzed.

Basic Principles of Transcription and Translation: Genes provide the instructions for making specific proteins. But a gene does not build a protein directly. The bridge between DNA and protein synthesis is the nucleic acid RNA. RNA is chemically similar to DNA except that it contains ribose instead of deoxyribose as its sugar and has the nitrogenous base uracil rather than thymine (see Figure 5.23).

Thus, each nucleotide along a DNA strand has A, G, C, or T as its base, and each nucleotide along an RNA strand has A, G, C, or U as its base. An RNA molecule usually consists of a single strand.

In mammalian testes and ovaries, the two sex chromosomes segregate during meiosis. Each egg receives one X chromosome. In contrast, sperm fall into two categories: Half the sperm cells a male produces receive an X chromosome, and half receive a Y chromosome. We can trace the sex of each offspring to the events of conception: If a sperm cell bearing an X chromosome fertilizes an egg, the zygote is XX, a female; if a sperm cell containing a Y chromosome fertilizes an egg, the zygote is XY, a male (see Figure 15.6a).

Thus, in general, sex determination is a matter of chance—a fifty-fifty chance. Note that the mammalian X-Y system isn't the only chromosomal system for determining sex. Figure 15.6b-d illustrates three other systems

About one out of ten African-Americans have sickle-cell trait, an unusually high frequency of heterozygotes for an allele with severe detrimental effects in homozygotes. Why haven't evolutionary processes resulted in the disappearance of the allele among this population? One explanation is that having a single copy of the sickle-cell allele reduces the frequency and severity of malaria attacks, especially among young children. The malaria parasite spends part of its life cycle in red blood cells (see Figure 28.16), and the presence of even heterozygous amounts of sickle-cell hemoglobin results in lower parasite densities and hence reduced malaria symptoms.

Thus, in tropical Africa, where infection with the malaria parasite is common, the sickle-cell allele confers an advantage to heterozygotes even though it is harmful in the homozygous state. (The balance between these two effects will be discussed in Concept 23.4; see Make Connections Figure 23.18.) The relatively high frequency of African-Americans with sickle-cell trait is a vestige of their African ancestry.

At anaphase II, the release of sister chromatid cohesion at the centromeres allows the sister chromatids to separate.

Thus, sister chromatid cohesion and crossing over, acting together, play an essential role in the lining up of chromosomes by homologous pairs at metaphase I.

The occurrence of pairs of homologous chromosomes in each human somatic cell is a consequence of our sexual origins. We inherit one chromosome of a pair from each parent.

Thus, the 46 chromosomes in our somatic cells are actually two sets of 23 chromosomes—a maternal set (from our mother) and a paternal set (from our father). The number of chromosomes in a single set is represented by n.

Just as natural selection favors bacteria that have receptors altered by mutation or that have enzymes that cut phage DNA, it also favors phage mutants that can bind to altered receptors or that are resistant to enzymes.

Thus, the bacterium-phage relationship is in constant evolutionary flux.

If the amount of glucose in the cell increases, the cAMP concentration falls, and without cAMP, CRP detaches from the lac operon. Because CRP is inactive, RNA polymerase binds less efficiently to the promoter, and transcription of the lac operon proceeds only at a low level, even when lactose is present (Figure 18.5b).

Thus, the lac operon is under dual control: negative control by the lac repressor and positive control by CRP. The state of the lac repressor (with allolactose bound or without it) determines whether or not transcription of the lac operon's genes occurs at all; the state of CRP (with bound cAMP or without it) controls the rate of transcription if the operon is repressor-free. It is as though the operon has both an on-off switch and a volume control.

Operons: The Basic Concept E. coli synthesizes the amino acid tryptophan from a precursor molecule in the three-step pathway shown in Figure 18.2.Each reaction in the pathway is catalyzed by a specific enzyme, and the five genes that code for the subunits of these enzymes are clustered together on the bacterial chromosome. A single promoter serves all five genes, which together constitute a transcription unit. (Recall that a promoter is a site where RNA polymerase can bind to DNA and begin transcription; see Figure 17.8.)

Thus, transcription gives rise to one long mRNA molecule that codes for the five polypeptides making up the enzymes in the tryptophan pathway (Figure 18.3a). The cell can translate this one mRNA into five separate polypeptides because the mRNA is punctuated with start and stop codons that signal where the coding sequence for each polypeptide begins and ends

Following enzymatic degradation of the mRNA, a second DNA strand, complementary to the first, is synthesized by DNA polymerase. The resulting double-stranded DNA is called complementary DNA (cDNA). (Made from mRNA, cDNA lacks introns and can be used for protein expression in bacteria, as mentioned earlier.)

To analyze the timing of expression of the Drosophila gene of interest, for example, we would first isolate all the mRNAs from different stages of Drosophila embryos and make cDNA from each stage (Figure 20.11).

. Using Mendelian symbols, our cross is PpYyRr * Ppyyrr. What fraction of offspring from this cross are predicted to exhibit the recessive phenotypes for at least two of the three characters?

To answer this question, we can start by listing all genotypes we could get that fulfill this condition: ppyyRr, ppYyrr, Ppyyrr, PPyyrr, and ppyyrr. (Because the condition is at least two recessive traits, it includes the last genotype, which shows all three recessive traits.) Next, we calculate the probability for each of these genotypes resulting from our PpYyRr * Ppyyrr cross by multiplying together the individual probabilities for the allele pairs, just as we did in our dihybrid example.

Finally, the length of time each protein functions in the cell is strictly regulated by selective degradation. Many proteins, such as the cyclins involved in regulating the cell cycle, must be relatively short-lived if the cell is to function appropriately (see Figure 12.16).

To mark a protein for destruction, the cell commonly attaches molecules of a small protein called ubiquitin to the protein. Giant protein complexes called proteasomes then recognize the ubiquitintagged proteins and degrade them.

Biologists use DNA technology to study gene expression and function:

To see how a biological system works, scientists seek to understand the functioning of the system's component parts. Analysis of when and where a gene or group of genes is expressed can provide important clues about their function.

To check the recombinant plasmids after they have been copied many times in host cells (see Figure 20.4), a researcher might cut the products again using the same restriction enzyme, expecting two DNA fragments, one the size of the plasmid and one the size of the inserted DNA

To separate and visualize the fragments, researchers carry out a technique called gel electrophoresis, which uses a gel made of a polymer as a molecular sieve to separate out a mixture of nucleic acid fragments by length (Figure 20.6). Gel electrophoresis is used in conjunction with many different techniques in molecular biology. Now that we have discussed the cloning vector in some detail, let's consider the foreign DNA to be inserted. The most common way to obtain many copies of the gene to be cloned is by PCR, described next

As Figure 15.9 shows, both of the combinations of traits not seen in the P generation (called nonparental phenotypes) were also produced in Morgan's experiments, suggesting that the body color and wing size alleles are not always linked genetically.

To understand this conclusion, we need to further explore genetic recombination, the production of offspring with combinations of traits that differ from those found in either P generation parent.

Capsids and Envelopes: The protein shell enclosing the viral genome is called a capsid. Depending on the type of virus, the capsid may be rod-shaped, polyhedral, or more complex in shape. Capsids are built from a large number of protein subunits called capsomeres, but the number of different kinds of proteins in a capsid is usually small.

Tobacco mosaic virus has a rigid, rod-shaped capsid made from over 1,000 molecules of a single type of protein arranged in a helix; rod-shaped viruses are commonly called helical viruses for this reason (Figure 19.3a). Adenoviruses, which infect the respiratory tracts of animals, have 252 identical protein molecules arranged in a polyhedral capsid with 20 triangular facets—an icosahedron; thus, these and other similarly shaped viruses are referred to as icosahedral viruses (Figure 19.3b).

DNA is the genetic material:

Today, even schoolchildren have heard of DNA, and scientists routinely manipulate DNA in the laboratory. Early in the 20th century, however, identifying the molecules of inheritance loomed as a major challenge to biologists.

Other kinds of signal peptides are used to target polypeptides to mitochondria, chloroplasts, the interior of the nucleus, and other organelles that are not part of the endomembrane system. The critical difference in these cases is that translation is completed in the cytosol before the polypeptide is imported into the organelle.

Translocation mechanisms also vary, but in all cases studied to date, the "postal zip codes" that address proteins for secretion or to cellular locations are signal peptides of some sort. Bacteria also employ signal peptides to target proteins to the plasma membrane for secretion.

Candidates for the original sources of viral genomes include plasmids and transposons. Plasmids are small, circular DNA molecules found in bacteria and in the unicellular eukaryotes called yeasts. Plasmids exist apart from and can replicate independently of the bacterial chromosome and are occasionally transferred between cells.

Transposons are DNA segments that can move from one location to another within a cell's genome. Thus, plasmids, transposons, and viruses all share an important feature: They are mobile genetic elements. (We'll discuss plasmids in more detail in Concepts 20.1 and 27.2 and transposons in Concept 21.4.)

Eventually, however, anthropological investigations ferreted out how the disease was spread: ritual cannibalism, a widespread practice among South Fore natives at that time.

Two characteristics of prions are especially alarming. First, prions act very slowly, with an incubation period of at least ten years before symptoms develop. The lengthy incubation period prevents sources of infection from being identified until long after the first cases appear, allowing many more infections to occur. Second, prions are virtually indestructible; they are not destroyed or deactivated by heating to normal cooking temperatures. To date, there is no known cure for prion diseases, and the only hope for developing effective treatments lies in understanding the process of infection.

Cytoplasmic Determinants and Inductive Signals: What generates the first differences among cells in an early embryo? And what controls the differentiation of all the various cell types as development proceeds? By this point in the chapter, you can probably deduce the answer: The specific genes expressed in any particular cell of a developing organism determine its path.

Two sources of information, used to varying extents in different species, "tell" a cell which genes to express at any given time during embryonic development. One important source of information early in development is the egg's cytoplasm, which contains both RNA and proteins encoded by the mother's DNA. The cytoplasm of an unfertilized egg is not homogeneous. Messenger RNA, proteins, other substances, and organelles are distributed unevenly in the unfertilized egg, and this unevenness has a profound impact on the development of the future embryo in many species. Maternal substances in the egg that influence the course of early development are called cytoplasmic determinants (Figure 18.17a). After fertilization, early mitotic divisions distribute the zygote's cytoplasm into separate cells. The nuclei of these cells may thus be exposed to different cytoplasmic determinants, depending on which portions of the zygotic cytoplasm a cell received. The combination of cytoplasmic determinants in a cell helps determine its developmental fate by regulating expression of the cell's genes during the course of cell differentiation

A program of differential gene expression leads to the different cell types in a multicellular organism: In the embryonic development of multicellular organisms, a fertilized egg (a zygote) gives rise to cells of many different types, each with a different structure and corresponding function.

Typically, cells are organized into tissues, tissues into organs, organs into organ systems, and organ systems into the whole organism. Thus, any developmental program must produce cells of different types that form higher-level structures arranged in a particular way in three dimensions. The processes that occur during development in plants and animals are detailed in Chapters 35 and 47, respectively. In this chapter, we focus on the program of regulation of gene expression that orchestrates development, using a few animal species as examples.

The two chromosomes referred to as X and Y are an important exception to the general pattern of homologous chromosomes in human somatic cells.

Typically, human females have a homologous pair of X chromosomes (XX), while males have one X and one Y chromosome (XY; see Figure 13.3). Only small parts of the X and Y are homologous.

The probability for the other possible way— the recessive allele from the egg and the dominant allele from the sperm—is also 1 ⁄ 4 (see Figure 14.9).

Using the rule of addition, then, we can calculate the probability of an F2 heterozygote as 1 ⁄ 4 + 1 ⁄ 4 = 1 ⁄ 2.

A recently discovered class of small ncRNAs is called piwi-interacting RNAs, or piRNAs. These RNAs also induce formation of heterochromatin, blocking expression of some parasitic DNA elements in the genome known as transposons. (Transposons are discussed in Concepts 21.4 and 21.5.)

Usually 24-31 nucleotides in length, piRNAs are processed from a longer, single-stranded RNA precursor. They play an indispensable role in the germ cells of many animal species, where they appear to help reestablish appropriate methylation patterns in the genome during gamete formation.

Recently, scientists have become concerned about the possible effects of climate change on worldwide viral transmission. Dengue fever, also mosquito-borne, has appeared in Florida and Portugal, regions where it had not been seen before. The possibility that global climate change has allowed mosquito species carrying these viruses to expand their ranges and interact more is troubling because of the increased chance of a mutation allowing a virus species to jump to a new host. This is an area of active research by scientists applying climate change models to what is known about the habitat requirements of mosquito species.

Viral Diseases in Plants: More than 2,000 types of viral diseases of plants are known, and together they account for an estimated annual loss of $15 billion worldwide due to their destruction of agricultural and horticultural crops. Common signs of viral infection include bleached or brown spots on leaves and fruits (Figure 19.12), stunted growth, and damaged flowers or roots, all of which can diminish the yield and quality of crops.

Plant viruses have the same basic structure and mode of replication as animal viruses. Most plant viruses discovered thus far, including tobacco mosaic virus (TMV), have an RNA genome. Many have a helical capsid, like TMV, while others have an icosahedral capsid (see Figure 19.3b).

Viral diseases of plants spread by two major routes. In the first route, called horizontal transmission, a plant is infected from an external source of the virus. Because the invading virus must get past the plant's outer protective layer of cells (the epidermis), a plant becomes more susceptible to viral infections if it has been damaged by wind, injury, or herbivores.

Different strains of influenza A are given standardized names; for example, both the strain that caused the 1918 flu and the one that caused the 2009 pandemic flu are called H1N1. The name identifies which forms of two viral surface proteins are present: hemagglutinin (HA) and neuraminidase (NA). There are 16 different types of hemagglutinin, a protein that helps the flu virus attach to host cells, and 9 types of neuraminidase, an enzyme that helps release new virus particles from infected cells.

Waterbirds have been found that carry viruses with all possible combinations of HA and NA. Variations of the hemagglutinin protein are used each year to generate vaccines against the strains predicted most likely to occur the next year.

Why is this requirement inconsistent with like-with-like pairing of bases? Adenine and guanine are purines, nitrogenous bases with two organic rings, while cytosine and thymine are nitrogenous bases called pyrimidines, which have a single ring. Pairing a purine with a pyrimidine is the only combination that results in a uniform diameter for the double helix:

Watson and Crick reasoned that there must be additional specificity of pairing dictated by the structure of the bases. Each base has chemical side groups that can form hydrogen bonds with its appropriate partner: Adenine can form two hydrogen bonds with thymine and only thymine; guanine forms three hydrogen bonds with cytosine and only cytosine. In shorthand, A pairs with T, and G pairs with C (Figure 16.8)

Watson and Crick began building models of a double helix that would conform to the X-ray measurements and what was then known about the chemistry of DNA, including Chargaff's rule of base equivalences. Having also read an unpublished annual report summarizing Franklin's work, they knew she had concluded that the sugar-phosphate backbones were on the outside of the DNA molecule, contrary to their working model. Franklin's arrangement was appealing because it put the negatively charged phosphate groups facing the aqueous surroundings, while the relatively hydrophobic nitrogenous bases were hidden in the interior.

Watson constructed such a model, shown in the small photo on the first page of this chapter. In this model, the two sugar-phosphate backbones are antiparallel—that is, their subunits run in opposite directions (see Figure 16.7). You can imagine the overall arrangement as a rope ladder with rigid rungs. The side ropes represent the sugar-phosphate backbones, and the rungs represent pairs of nitrogenous bases. Now imagine twisting the ladder to form a helix. Franklin's X-ray data indicated that the helix makes one full turn every 3.4 nm along its length. With the bases stacked just 0.34 nm apart, there are ten layers of base pairs, or rungs of the ladder, in each full turn of the helix.

heredity

We all know that offspring resemble their parents more than they do unrelated individuals. If you examine the family members shown in Figure 13.1, you can pick out some similar features among them. The transmission of traits from one generation to the next is called inheritance, or heredity (from the Latin heres, heir). However, sons and daughters are not identical copies of either parent or of their siblings.

The presence of introns in a gene may facilitate the evolution of new and potentially beneficial proteins as a result of a process known as exon shuffling (see Figure 21.16). Introns increase the probability of crossing over between the exons of alleles of a gene—simply by providing more terrain for crossovers without interrupting coding sequences. This might result in new combinations of exons and proteins with altered structure and function.

We can also imagine the occasional mixing and matching of exons between completely different (nonallelic) genes. Exon shuffling of either sort could lead to new proteins with novel combinations of functions. While most of the shuffling would result in nonbeneficial changes, occasionally a beneficial variant might arise.

What is the probability that the child will be unable to taste PTC?

We can also treat this as a monohybrid cross (Tt * Tt), but this time we want to know the chance that the offspring will be homozygous recessive (tt). That probability is 1 ⁄ 4. Finally, what is the chance that the child will have a widow's peak and be unable to taste PTC? Assuming that the genes for these two characters are on different chromosomes, the two pairs of alleles will assort independently in this dihybrid cross (WwTt * WwTt). Therefore, we can use the multiplication rule: 3 ⁄ 4 (chance of widow's peak) * 1 ⁄ 4 (chance of inability to taste PTC) = 3 ⁄ 16 (chance of widow's peak and inability to taste PTC).

Building a Polypeptide:

We can divide translation, the synthesis of a polypeptide, into three stages: initiation, elongation, and termination. All three require protein "factors" that aid in the translation process. Some steps of initiation and elongation also require energy, provided by the hydrolysis of guanosine triphosphate (GTP)

Consider the dihybrid cross between YyRr heterozygotes shown in Figure 14.8. We will focus first on the seed-color character. For a monohybrid cross of Yy plants, we can use a simple Punnett square to determine that the probabilities of the offspring genotypes are 1 ⁄ 4 for YY, 1 ⁄ 2 for Yy, and 1 ⁄ 4 for yy.

We can draw a second Punnett square to determine that the same probabilities apply to the offspring genotypes for seed shape: 1 ⁄ 4 RR, 1 ⁄ 2 Rr, and 1 ⁄ 4 rr. Knowing these probabilities, we can simply use the multiplication rule to determine the probability of each of the genotypes in the F2 generation.

Tossing a coin illustrates an important lesson about probability. For every toss, the probability of heads is 1 ⁄ 2. The outcome of any particular toss is unaffected by what has happened on previous trials.

We refer to phenomena such as coin tosses as independent events. Each toss of a coin, whether done sequentially with one coin or simultaneously with many, is independent of every other toss.

What Is a Gene? Revisiting the Question: Our definition of a gene has evolved over the past few chapters, as it has through the history of genetics. We began with the Mendelian concept of a gene as a discrete unit of inheritance that affects a phenotypic character (Chapter 14). We saw that Morgan and his colleagues assigned such genes to specific loci on chromosomes (Chapter 15).

We went on to view a gene as a region of specific nucleotide sequence along the length of the DNA molecule of a chromosome (Chapter 16). Finally, in this chapter, we have considered a functional definition of a gene as a DNA sequence that codes for a specific polypeptide chain or a functional RNA molecule, such as a tRNA. All these definitions are useful, depending on the context in which genes are being studied.

Interference with Normal Cell-Signaling Pathways The proteins encoded by many proto-oncogenes and tumorsuppressor genes are components of cell-signaling pathways. Let's take a closer look at how such proteins function in normal cells and what goes wrong with their function in cancer cells.

We will focus on the products of two key genes, the ras proto-oncogene and the p53 tumor-suppressor gene. Mutations in ras occur in about 30% of human cancers, and mutations in p53 in more than 50%

And like two separate coin tosses, the alleles of one gene segregate into gametes independently of another gene's alleles (the law of independent assortment).

We'll now look at two basic rules of probability that help us predict the outcome of the fusion of such gametes in simple monohybrid crosses and more complicated crosses as well.

Some of these genes code for proteins involved in translation, DNA repair, protein folding, and polysaccharide synthesis. Whether mimivirus evolved before the first cells and then developed an exploitative relationship with them or evolved more recently and simply scavenged genes from its hosts is not yet settled. Since 2013 several even larger viruses have been discovered that cannot be classified with any existing known virus. One such virus is 1 µm (1,000 nm) in diameter, with a dsDNA genome of around 2-2.5 Mb, larger than that of some small eukaryotes.

What's more, over 90% of its 2,000 or so genes are unrelated to cellular genes, inspiring the name it was given, pandoravirus. A second virus, called Pithovirus sibericum, with a diameter of 1.5 μm and 500 genes, was discovered in permanently frozen soil in Siberia. This virus, once thawed, was able to infect an amoeba after being frozen for 30,000 years! How these and all other viruses fit in the tree of life is an intriguing, unresolved question.

Figure 16.9 illustrates Watson and Crick's basic idea. To make it easier to follow, we show only a short section of double helix in untwisted form. Notice that if you cover one of the two DNA strands of Figure 16.9a, you can still determine its linear sequence of nucleotides by referring to the uncovered strand and applying the base-pairing rules. The two strands are complementary; each stores the information necessary to reconstruct the other.

When a cell copies a DNA molecule, each strand serves as a template for ordering nucleotides into a new, complementary strand. Nucleotides line up along the template strand according to the base-pairing rules and are linked to form the new strands. Where there was one double-stranded DNA molecule at the beginning of the process, there are soon two, each an exact replica of the "parental" molecule. The copying mechanism is analogous to using a photographic negative to make a positive image, which can in turn be used to make another negative, and so on.

The function of any cell, whether a single-celled eukaryote or a particular cell type in a multicellular organism, depends on the appropriate set of genes being expressed. The transcription factors of a cell must locate the right genes at the right time, a task on a par with finding a needle in a haystack.

When gene expression proceeds abnormally, serious imbalances and diseases, including cancer, can arise.

Positive Gene Regulation:

When glucose and lactose are both present in its environment, E. coli preferentially uses glucose. The enzymes for glucose breakdown in glycolysis (see Figure 9.9) are continually present. Only when lactose is present and glucose is in short supply does E. coli use lactose as an energy source, and only then does it synthesize appreciable quantities of the enzymes for lactose breakdown.

How can a protein, which cannot replicate itself, be a transmissible pathogen? According to the leading model, a prion is a misfolded form of a protein normally present in brain cells.

When the prion gets into a cell containing the normal form of the protein, the prion somehow converts normal protein molecules to the misfolded prion versions. Several prions then aggregate into a complex that can convert other normal proteins to prions, which join the chain (Figure 19.13).

Next in RT-PCR is the PCR step (see Figure 20.7). As you will recall, PCR is a way of rapidly making many copies of one specific stretch of double-stranded DNA, using primers that hybridize to the opposite ends of the segment of interest. In our case, we would add primers corresponding to a segment of our Drosophila gene, using the cDNA from each embryonic stage as a template for PCR amplification in separate samples

When the products are analyzed on a gel, copies of the amplified region will be observed as bands only in samples that originally contained mRNA from the gene of interest. An enhancement called quantitative RT-PCR (qRT-PCR) uses a fluorescent dye that fluoresces only when bound to a doublestranded PCR product. The newer quantitative PCR machines can detect the light and measure the PCR product, thus avoiding the need for electrophoresis while also providing quantitative data, a distinct advantage. RT-PCR or qRT-PCR can also be carried out with mRNAs collected from different tissues at one time to discover which tissue is producing a specific mRNA.

Replicative Cycles of Animal Viruses: Everyone has suffered from viral infections, whether cold sores, influenza, or the common cold. Like all viruses, those that cause illness in humans and other animals can replicate only inside host cells. Many variations on the basic scheme of viral infection and replication are represented among the animal viruses. One key variable is the nature of the viral genome (double- or single-stranded DNA or RNA). Another variable is the presence or absence of a membranous envelope.

Whereas few bacteriophages have an envelope or RNA genome, many animal viruses have both. In fact, nearly all animal viruses with RNA genomes have an envelope, as do some with DNA genomes. Rather than consider all the mechanisms of viral infection and replication, we will focus first on the roles of viral envelopes and then on the functioning of RNA as the genetic material of many animal viruses.

The brief but celebrated partnership that solved the puzzle of DNA structure began soon after Watson journeyed to Cambridge University, where Crick was studying protein structure with a technique called X-ray crystallography (see Figure 5.21).

While visiting the laboratory of Maurice Wilkins, Watson saw an X-ray diffraction image of DNA produced by Wilkins's accomplished colleague Rosalind Franklin (Figure 16.6). Images produced by X-ray crystallography are not actually pictures of molecules. The spots and smudges in the image were produced by X-rays that were diffracted (deflected) as they passed through aligned fibers of purified DNA.

Morgan's first mutant.:

Wild-type Drosophila flies have red eyes (left). Among his flies, Morgan discovered a mutant male with white eyes (right). This variation made it possible for Morgan to trace a gene for eye color to a specific chromosome.

Amplifying DNA: The Polymerase Chain Reaction (PCR) and Its Use in DNA Cloning: Today, most researchers have some information about the sequence of the gene or other DNA segment they want to clone. Using this information, they can start with the entire collection of genomic DNA from the particular species of interest and obtain many copies of the desired gene by using a technique called the polymerase chain reaction, or PCR. Figure 20.7 illustrates the steps in PCR.

Within a few hours, this technique can make billions of copies of a specific target DNA segment in a sample, even if that segment makes up less than 0.001% of the total DNA in the sample. In the PCR procedure, a three-step cycle brings about a chain reaction that produces an exponentially growing population of identical DNA molecules. During each cycle, the reaction mixture is heated to denature (separate) the strands of the double-stranded DNA and then cooled to allow annealing (hydrogen bonding) of short, single-stranded DNA primers complementary to sequences on opposite strands at each end of the target sequence; finally, a heat-stable DNA polymerase extends the primers in the 5¿ S 3¿ direction. If a standard DNA polymerase were used, the protein would be denatured along with the DNA during the first heating step and would have to be replaced after each cycle. The key to automating PCR was the discovery of an unusual heat-stable DNA polymerase called Taq polymerase, named after the bacterial species from which it was first isolated. This bacterial species, Thermus aquaticus, lives in hot springs, and the stability of its DNA polymerase at high temperatures is an evolutionary adaptation that enables the enzyme to function at temperatures up to 95°C.

In the Problem-Solving Exercise, you'll work with a few common single nucleotide-pair substitution mutations in the gene encoding insulin, some or all of which may lead to diabetes.

You will classify these mutations into one of the types we just described and characterize the change in amino acid sequence.

How do such viruses burst on the human scene, giving rise to harmful diseases that were previously rare or even unknown? Three processes contribute to the emergence of viral diseases. The first, and perhaps most important, is the mutation of existing viruses. RNA viruses tend to have an unusually high rate of mutation because viral RNA polymerases do not proofread and correct errors in replicating their RNA genomes. Some mutations change existing viruses into new genetic varieties (strains) that can cause disease, even in individuals who are immune to the ancestral virus. For instance, seasonal flu epidemics are caused by new strains of influenza virus genetically different enough from earlier strains that people have little immunity to them.

You'll see an example of this process in the Scientific Skills Exercise, where you'll analyze genetic changes in variants of the H1N1 flu virus and correlate them with spread of the disease. A second process that can lead to the emergence of viral diseases is the dissemination of a viral disease from a small, isolated human population. For instance, AIDS went unnamed and virtually unnoticed for decades before it began to spread around the world. In this case, technological and social factors, including affordable international travel, blood transfusions, sexual promiscuity, and the abuse of intravenous drugs, allowed a previously rare human disease to become a global scourge.

Down Syndrome (Trisomy 21): One aneuploid condition, Down syndrome, affects approximately one out of every 830 children born in the United States (Figure 15.15). Down syndrome is usually the result of an extra chromosome 21, so that each body cell has a total of 47 chromosomes. Because the cells are trisomic for chromosome 21, Down syndrome is often called trisomy 21. Down syndrome includes characteristic facial features, short stature, correctable heart defects, and developmental delays.

ndividuals with Down syndrome have an increased chance of developing leukemia and Alzheimer's disease but have a lower rate of high blood pressure, atherosclerosis (hardening of the arteries), stroke, and many types of solid tumors. Although people with Down syndrome, on average, have a life span shorter than normal, most, with proper medical treatment, live to middle age and beyond. Many live independently or at home with their families, are employed, and are valuable contributors to their communities. Almost all males and about half of females with Down syndrome are sexually underdeveloped and sterile.

Prophase II

• A spindle apparatus forms. * In late prophase II (not shown here), chromosomes, each still composed of two chromatids associated at the centromere, are moved by microtubules toward the metaphase II plate.

Anaphase II: Sister chromatids separate

• Breakdown of proteins holding the sister chromatids together at the centromere allows the chromatids to separate. The chromatids move toward opposite poles as individual chromosomes.

Anaphase I: Sister chromatids remain attached Homologous chromosomes separate The two homologous chromosomes of each pair separate.

• Breakdown of proteins that are responsible for sister chromatid cohesion along chromatid arms allows homologs to separate. • The homologs move toward opposite poles, guided by the spindle apparatus. • Sister chromatid cohesion persists at the centromere, causing chromatids to move as a unit toward the same pole.

Prophase I : Centrosome (with centriole pair) Chiasmata Sister chromatids Spindle microtubules Fragments of nuclear envelope Pair of homologous chromosomes Centromere Fragments of nuclear envelope Duplicated homologous chromosomes (red and blue) pair up and exchange segments; 2n = 6 in this example.

• Centrosome movement, spindle formation, and nuclear envelope breakdown occur as in mitosis. Chromosomes condense progressively throughout prophase I. • During early prophase I, before the stage shown above, each chromosome pairs with its homolog, aligned gene by gene, and crossing over occurs: The DNA molecules of nonsister chromatids are broken (by proteins) and are rejoined to each other. • At the stage shown above, each homologous pair has one or more X-shaped regions called chiasmata (singular, chiasma), where crossovers have occurred.

Telophase II and Cytokinesis: Haploid daughter cells forming

• Nuclei form, the chromosomes begin decondensing, and cytokinesis occurs. • The meiotic division of one parent cell produces four daughter cells, each with a haploid set of (unduplicated) chromosomes. • The four daughter cells are genetically distinct from one another and from the parent cell.

Metaphase I Kinetochore (at centromere) Kinetochore microtubules Metaphase plate Chromosomes line up by homologous pairs.

• Pairs of homologous chromosomes are now arranged at the metaphase plate, with one chromosome of each pair facing each pole. • Both chromatids of one homolog are attached to kinetochore microtubules from one pole; the chromatids of the other homolog are attached to microtubules from the opposite pole.

Metaphase II

• The chromosomes are positioned at the metaphase plate as in mitosis. • Because of crossing over in meiosis I, the two sister chromatids of each chromosome are not genetically identical. • The kinetochores of sister chromatids are attached to microtubules extending from opposite poles.

Telophase I and Cytokinesis :Two haploid cells form; each chromosome still consists of two sister chromatids. Cleavage furrow

• When telophase I begins, each half of the cell has a complete haploid set of duplicated chromosomes. Each chromosome is composed of two sister chromatids; one or both chromatids include regions of nonsister chromatid DNA. • Cytokinesis (division of the cytoplasm) usually occurs simultaneously with telophase I, forming two haploid daughter cells. • In animal cells like these, a cleavage furrow forms. (In plant cells, a cell plate forms.) • In some species, chromosomes decondense and nuclear envelopes form. • No chromosome duplication occurs between meiosis I and meiosis II.