Chapter 8

What is a mutagen?

A mutagen is a substance that causes increased mutation rate. A substance that can cause a mutation A substance in the environment that directly or indirectly causes mutation A mutagen is a substance that has the ability to induce mutations

What is an F+ cell?

An F+ cell had the fertility factor, a plasmid which allows the cell to produce the genes for horizontal gene transfer. A cell that carries the fertility factor within its plasmids

What is an operon?

An operon is all the genes related to one purpose, under one promoter. A cluster of genes that code for a set of enzymes or proteins. The operon is the section of a genome that codes for a certain enzyme or protein, including both the structural genes and the control region.

Why is base pairing in DNA important?

Any mistake in the base pairing leads to mutation, which can be devastating or be the driving force of evolution. It determines the specific function of one segment of DNA The base pairing is the code for all amino acid and protein synthesis. If the base pairs get out of order, the protein could be affected and therefore can affect the function of the cell. DNA base pairing is important because it ensures that when a cell replicates, each daughter cell will have at least one correctly functioning strand of DNA

How would you isolate an antibiotic-resistant bacterium? An antibiotic-sensitive bacterium?

Culture the bacteria on a plate with antibiotics. Those that grow are resistant. Replica plating is also useful to determine auxotrophic bacteria. By inoculating it onto an antibiotic present plate. Replica plating is used to put the bacteria onto different plates with different antimicrobial enviornments Resistant- by inoculating a plate containing the antibiotic Sensitive- by replica plating onto a medium with the antibiotic and a medium without the antibiotic Antibiotic-resistant: Inoculate the bacterium and plate it on a plate that contains the antibiotic. Antibiotic-sensitive: use replica plating. The bacterium should be spread on a "master" plate and then picked up by velvet or a nylon filter. They are then "stamped" onto other plates containing different antibiotics or no antibiotics and allowed to grow.

What type of enzyme breaks the DNA? What enzyme rejoins the pieces of DNA?

DNA Helicase breaks it, DNA Ligase rejoins. Restriction enzymes. DNA polymerase joins them Endonucleases break DNA and ligase joins pieces of DNA Breaks down: Restriction enzymes Rejoin: DNA ligase

Describe DNA replication, including the functions of DNA gyrase, DNA ligase, and DNA polymerase.

DNA gyrase relaxes the supercoiling of DNA. Helicase unzips the DNA strand, separating the double helix. DNA polymerase copies the corresponding nucleotide on the leading strand. RNA primers synthesizes the lagging strand discontinuously. DNA polymerase then digests the RNA primer and replaces it with DNA. DNA Ligase joints the fragments of the lagging strand. One strand of DNA is separated and replicated into two identical strands. To start gyrase relaxes the strand of and helicase then unwinds the strand. Then Free nucleotides line up with their appropriate bases and are joined via DNA polymerase (should be ligase not polymerase) In DNA replication, a strand of DNA is relaxed or loosened by DNA gyrase and "unzipped" by DNA helicase. The partial DNA strands are proofread by DNA polymerase while the complementary base pairs are attached by DNA ligase. DNA gyrase begins the process by relaxing the wound chromosome, and helicase fully unwinds it. The chromosome is separated and free nucleotides in the cytoplasm pair with the exposed bases. DNA polymerase joins the newly aligned pair to the overall strand, then the DNA ligase binds the new DNA strand to the old one.

How can mutations be repaired?

DNA ligase helps to repair DNA, as does DNA polymerase. Thymine dimers are fixed with photolyases. Nucleotide excision repair cuts out the incorrect bases. Excision repair and phagocytosis that break apart thymine dymers Excision repair mechanisms are one way that mutations can be fixed. Another way is Photolyases which breaks apart thymine dimers. Using photolyases, which use energy from visible light to separate cross-linked bases, or using nucleotide excision repair molecules, which remove the incorrect base and replace it with the correct base

What type of enzyme cuts the donor DNA?

Endonuclease The endonuclease

How do excision repair enzymes "know" which strand is incorrect?

Enzymes add a methyl group to some bases when DNA is made to track the one without methyl. The enzyme methylase adds methyl groups to correctly paired base pairs. The incorrect ones are then distinguished Methylases bond to the correct strand so that the endonuclease cuts the other strand Methylase follows behind the DNA polymerase and adds a methyl group to the correctly paired base pairs. The incorrect pairs do not receive a methyl group and are therefore distinguishable from the correct ones.

Compare conjugation between the following pairs: F+ X F-, Hfr X F-.

F+ X F- = The F+ will pass on the plasmid to the F-, making it F+. Hfr X F- = The Hfr will pass on a portion of the chromosome to the F-, making it a recombinant F- cell. F+ x F--: the F+ gives one strand of its plasmid to the F-- F-- x Hfr: the Hfr replicates its entire F+ containing chromosome. The F factor leads the strand to the F-- cell. The strand usually breaks before the whole chromosome is transferred, resulting in the F-- cell remaining F-- because it does not get the entire gene Conjugation between F+ X F- involves the transfer of a copy of the F plasmid from the + to the - microbe. In Hfr X F- conjugation, the whole plasmid and all genomic materials are transferred to the lacking cell.

When both glucose and lactose are present, why will cells use glucose First?

Glucose produces more energy than lactose, so it is more efficient to use it first. Glucose offers more energy and its presence prevents the operons that metabolize lactose from functioning Glucose is the most effective carbon source for cells, and it prevents cAMP buildup In order to use lactose as an energy source, the cell must produce cyclic AMP. cAMP is only produced when the environment is glucose deficient. Glucose yields more energy than lactose does so many cells will use all the glucose in their environment before switching to lactose.

Why can translation begin before transcription is complete in prokaryotes but not in eukaryotes?

In eukaryotes, there are no ribosomes in the nucleus, so they have to wait for the genetic information to exit the nucleus. The ribosomes are not separated by a nucleus in prokaryotic cells In prokaryotes, transcription occurs in the cytoplasm where the ribosomes needed for translation are already present, so as soon as the UAG codon is available, translation begins. Because in eukaryotic cells, ribosomes are located outside of a membrane bound nucleus whereas in prokaryotic cells, ribosomes are not separated by a membrane.

What is the principle behind the Ames test?

It is based on the fact that mutations can be reverted, and tests the speed at which they are reverted compared to the spontaneous reversion rate. The amount of mutation present before a mutation is related to the number of bacteria after exposure to the mutagen Exposing mutant microbes to mutagens will cause a reversion or a second mutation that reverses the effect of the first The principle behind the Ames Test is that when a sample is exposed to a mutagen, the auxotrophic strain reverts back to a histidine synthesizing strain. Quantitatively, the level of mutation that was present before the mutagen is directly related to the number of colonies arising after the mutagen exposure.

What is the role of cAMP in catabolite repression?

It requires the absence of glucose and the presence of lactose. It binds to CAP, which binds to the lac promoter, and allow transcription to begin. cAMP levels within the cell are low and don't allow for sugars or carbon sources other than glucose to be metabolized cAMP's role, once produced, is to bind to a catabolite activator protein and then bind to a promoter which will remove the lactose repressor (if lactose is present). Therefore, the cell switches from glucose to lactose as its energy source. cAMP buildup tells the cell that catabolite repression is no longer occurring, meaning the cell is short of glucose so other carbon sources should be used.

On what basis are missense, nonsense, and frameshift mutations distinguished?

Missense - One amino acid is changed. The rest remain the same. Nonsense - a premature stop codon is created. Frameshift - potentially changes every amino acid after the mutation. Missense- one base is substituted, resulting in an amino acid change Nonsense- one or more bases are substituted, resulting in a premature stop codon Frameshift- one or more bases are removed from or added to the sequence, resulting in the rest of the amino acids being incorrect They are distinguished by the amount of change they create. A missense changes a base pair to possibly result in a different amino acid. A nonsense changes a whole codon into a stop codon. A frameshift adds or removes base pairs which changes every codon after the point of shift because the codons are still read in threes.

How do mutagens affect the mutation rate?

Mutagens increase the mutation rate dramatically, from about 1/106 to 1/105-1/1 By increasing the chances of a mutation happening. Sometimes up to 100x They increase mutation rate by about 10x up to 1000x The introduction of a mutagen increases the mutation rate by 10-100 times

Do all mutagens cause cancer?

No, not all mutagens are carcinogenic. No, but most do and cancer tends to be a result of many mutagens acting together No No but more than 90% do. Most often, cancer is the result of multiple mutations which would require multiple mutagens.

Does a base substitution always result in a different amino acid?

No, some are silent and code for the same amino acid. No. Amino acids can be created even with the single substitutions No, because the code is redundant, meaning several combinations of nucleotides can code for the same amino acid No, Amino acids can be made from a variety of base pairs so a single base substitution could result in the same amino acid being produced.

What types of genes do plasmids carry?

Plasmids can carry many types of genes, some of which increase pathogenicity of a disease-causing microbe, allowing for rarer catabolic pathways and resistance to medicine. Genes that are non-essential to the cell under ordinary circumstances but are still needed from time to time. These genes can be conjugative or nonconjugative. Plasmids carry genes that are not necessary for cell survival but can increase the effect of the cell. For example: proteins to enhance pathogenicity, proteins to neutralize or denature antibiotics, and genes for the formation of sex pili.

Give a clinical application of genomics.

Predicting future strains of diseases Understanding the mechanism of diseases Identifying and testing for genetic markers Mapping the genome of bacteria and discovering the process that makes their proteins and stopping the DNA from doing that specific replication Genomics can be used to track diseases like the west nile virus Genomics can be used to better understand what combinations of base pairs code for specific diseases so that they can be removed

How can a mutation be beneficial?

Some mutations cause things like resistance to antibiotics, or other beneficial changes to metabolic activity. When a mutation changes the aspect of the cell that made it vulnerable Mutations can be beneficial by introducing new gene variation into the gene pool and also lead to the adaptation of the mutated individuals. By altering a protein to be stronger or have more beneficial properties than before

Natural selection means that the environment favors survival of some genotypes. From where does diversity in genotypes come?

The adaptation of organisms through natural selection is one way. Mutation is the driving force of evolution. Horizontal gene transfer also plays a major role, allowing microbes to spread genes. Diversity comes from mutation, conjugation, transposition, and recombination. Genotype diversity is a result of horizontal gene transfer of other microbial genes or the result of mutations.

How are mutations caused by chemicals? By radiation?

The chemicals are randomly incorporated into DNA and then cause mistakes during the building of the base pairs. In radiation, instead of altering the nucleotides directly, electrons and free radicals can cause damage, or ions can bond with bases. Chemicals: Chemical caused mutations are usually small changes to the DNA such as point-mutations. Radiation: Radiation caused mutations are usually larger changes to chromosomes Chemicals- either replace nitrogenous bases, resulting in pairing mutations, or inserting themselves between bases to cause frameshifts Radiation- either by ionizing electrons which form free radicals that bond incorrectly with bases or by interrupting bonds in the sugar-phosphate backbone

Why can't RNA transcript be used for translation?

The exons aren't removed yet. The introns must be removed first. They are the non-coding region Because the exons have not been removed yet, and it is a waste of energy to produce them. RNA transcript can't be used for translation because the exons have to be removed before translation can occur.

What is the role of the promoter, terminator, and mRNA in transcription?

The promoter is the binding site at the beginning of a gene, RNA polymerase will bind there.The terminator is the location where replication will stop. mRNA, messenger RNA, carries the genetic information from the DNA to the ribosomes. mRNA carries the coded information for making specific proteins from DNA to ribosomes. The promoter is a site on DNA where RNA polymerase can begin its binding. When RNA reaches the terminator, it concludes replication. mRNA is the carrier of the coded information for specific proteins from DNA to ribosomes. The promoter is the beginning site of RNA transcription while the terminator is the ending site. mRNA carries the genetic information for protein synthesis from DNA to ribosomes where the protein is made. The promoter is the site where the RNA polymerase binds to the DNA to start transcription. The terminator is the site on the DNA strand that tells the polymerase to stop transcription.

Will transcription of the lac operon occur in the presence of lactose and glucose? In the presence of lactose and the absence of glucose? In the presence of glucose and the absence of lactose?

Transcription of the lac operon will only occur in the presence of lactose but not glucose. No, a lac operon occurs when lactose is present and glucose is not. So, yes to the second, no to the third. Just lactose present- yes Just glucose present- no Glucose and lactose present- no Lactose and glucose present- No Just lactose present- Yes Just glucose present- No

What causes transcription of an inducible enzyme?

The repressor enzyme will not be attached to the operator. The deactivation or removal of a repressor molecule The presence of an inducer molecule The removal or deactivation of the repressor molecule will result in the transcription of an inducible enzyme.

What causes transcription of a repressible enzyme?

The repressor will be inactive, which is its standard state. When a compound is not present or lacking, the repressible enzyme will transcribe The absence of a repressor molecule A lack of or deficiency of a compound will lead to the transcription and expression of a repressible enzyme to makeup for the lacking compound. If the compound becomes available again, the transcription of the repressible enzyme will stop.

Why did encapsulated bacteria kill the mouse while nonencapsulated bacteria did not? What killed the mouse in (d)?

The unencapsulated bacteria were more easily killed by the immune system because they had less protection. The unencapsulated bacteria gained the genes for capsules through the horizontal gene transfer from dead bacteria. Encapsulated bacteria are not prone to phagocytosis (the mouse's immune system) Transfer of DNA allows non-encapsulated cells to "learn" how to encapsulate The encapsulated bacteria killed the mouse because the capsule protected the cells from phagocytosis, the mouse's main defense. In (d), the DNA from the heat-killed encapsulated cells transferred horizontally to the live, unencapsulated cells, which then learned how to survive phagocytosis and killed the mouse. The non-encapsulated

Differentiate between horizontal and vertical gene transfer.

Vertical is the passing on of genes from parent to offspring. Horizontal gene transfer is when a cell passes on genes to another cell in the same generation as itself. Vertical gene transfer is when genes are passed to offspring. Horizontal is when genes are transferred laterally to the same generation Vertical gene transfer occurs between parent and daughter cells, while horizontal gene transfer occurs between cells of the same generation Horizontal gene transfer occurs between bacteria and their surroundings/ "neighbors" when they exchange pieces of DNA. Vertical gene transfer is that of parent to offspring.

Why are R factors important in the treatment of infectious diseases?

They are dangerous because they allow for resistance to be shared, but can also be used by scientists to spread genes beneficial to people. They allow microbes to resistant to our typical treatment, so we must understand them to cure infection with microbes that have the R factor If the disease has an R factor, prescribing certain medications will do nothing because the bacteria have an r-determinate against those drugs, so alternate treatments must be found

Why are transposons sometimes referred to as "jumping genes"?

They can move from their position to another location in the DNA molecule. They frequently move from site to site on the same chromosome Because they can jump to different locations on the same chromosome or plasmid, or even to another plasmid or virus

Why do nucleotide analogue drugs and nitrous acid kill cells?

They change the DNA purposefully, so that all proteins from that DNA will be made incorrectly. They cause carcinogenic frame-shift mutations Nitrous acid converts adenine to pair with cytosine, destroying all replicated DNA and proteins. Analogue drugs function in the place of a nucleotide and cause random mistakes in base pairing The drugs add nucleoside analogs to the microbes which then replaces the purines with pyrimidines in the DNA. This can lead to carcinogenic frameshift mutations which lead to the death of the cell.

What is transduction?

Transduction is the forced transfer of bacterial DNA from one cell to another, facilitated by a bacteriophage who breaks down and accidentally takes the DNA, used in the next cells they infect. The transfer of DNA from one cell to another by a bacterio-phage Horizontal transfer of DNA packed inside of coated phages

How does conjugation differ from transformation?

Transformation is genes released into the environment, to be picked up at random, but conjugation requires the participation and interaction of two cells for a deliberate transfer of genes. Conjunction is mediated by one kind of plasmid. These genes transferred are non-essential Conjugation is horizontal transfer of DNA from plasmids while transformation is horizontal transfer of DNA from the chromosome

How does mRNA production in eukaryotes differ from the process in prokaryotes?

Translation can begin before transcription is done in prokaryotes. Transcription is done in the cytoplasm in prokaryotes. In eukaryotic cells, translation has to begin after transcription is done, and transcription occurs in the nucleus. In eukaryotic cells replication takes place in the nucleus In prokaryotes, the translation of mRNA can begin before transcription. In eukaryotes, translation can only occur once transcription is done and the mRNA has been moved through the nuclear membrane. In prokaryotes, mRNA is made using two RNA polymerases travelling in opposite directions around a single, circular chromosome. In eukaryotes, one RNA polymerase is used on each linear chromosome. Prokaryotes can also begin translation before transcription is complete, which eukaryotes cannot do. In prokaryotes, transcription occurs in the cytoplasm because there is no nucleus.

What is an auxotroph?

When an inability to produce a nutritional requirement naturally is present, a microbe is an auxotroph. A mutant microorganism with a nutritional requirement that is absent in the parent A mutant microbe that needs a nutrient to grow that its parent doesn't need A mutated organism that now needs a specific new nutrient to survive that the non-mutated organism did not need.