Genetics Exam 4, Practical Biology Chapter 13, Biology Final Exam, CHEM 527 Exam Past, CHEM 527 Exams
One major purpose of the protein called what is to tether replication components to the DNA, in particular to enhance the processivity of DNA polymerase
sliding clamp
Santa Gertrudis cattle have the combined traits of good beef production and hot weather tolerance as a result of selection and ____________
hybridization
DNA polymerase ensures that the correct nudlotide is incorporated into the growing strand of DNA by
monitoring the ability of the incoming nucleotide to form either an AT or GC bp
DNA polymerase ensures that the correct nucleotide is incorportated into the growing strand of DNA by
monitoring the ability of the incoming nucleotide to form either and AT of GC bp
An RNA molecule is a polymer composed of subunits known as ___.
nucleotides
Thymine, adenine, guanine, and cytosine are classified as
nucleotides
relationship between organisms in which both organisms benefit
mutualism
The two halves of a doubled chromosome structure are called
sister chromatids
slide 4 set 32 ????
-figured out
Which segment in Figure 13-6 will attach to genetic material with the sequence TCGA?
C
The DNA fragments synthesized in lagging strand is called
Okazaki fragment
Genomics
the study of genomes
___ is used to develop pure breeds.
Inbreeding
A _______is a small ring of DNA found in a bacterial cell
Plasmid
Wolves would be considered ___.
carnivores
Applications of Gene cloning: Recombinant DNA
-Biotechnology: the genetic alteration of animals, plants, and bacteria **Transgenic organisms: In eukaryotes the introduced DNA must insert into a chromosome to be retained and expressed.
Some Current Applications of Plant Biotechnology
2011 data: 90% of corn, soybean, and cotton crops are GMO -the method described above is used to make these crops
You can do Locus-specific PCR of a VNTR or a VNTR southern blot to analyze someones DNA profile (VNTR with a southern blot requires a bigger sample of DNA)
***Colin Pitchfork convicted in 1987 of rape and murder based on DNA evidence
Herbicide Resistance --- Glyphosate and others
-Broad spectrum herbicide -Blocks EPSP synthase, an enzyme in the pathway for synthesis of aromatic amino acids. -Animals lack this enzyme, so toxicity of glyphosate is low -Breaks down quickly in soil (1-2 weeks) -Cheap -Transgenic plants expressing an EPSP synthase gene from bacteria are resistant to glyphosate -Herbicide resistance - grow plants without chemicals is not realistic to support our whole population - spraying with organically synthesized compounds is always a risk, this particular compound is the least toxic we know of but not completely nontoxic, this kills all plants by blocking EPSP which blocks amino acid synthesis (all animals get amino acids from diet) glyphosate has low toxicity to all animals and it degrades quickly in soil -FDA put this on the carcinogen list SO it has the potential to cause cancer but it is not known to cause cancer, only people at risk are people who make or use this compound -need to spray less with a more toxic herbicide because it persists longer in the soil
Knockout Mice: Targeted DNA Integration Via recombination In Embryonic Stem Cells
*Don't need to be worry about this but be aware -direct DNA into specific sites in mice and only in mice -done by homologous recombination -Create plasmid w seqeunces that are homologous to sites in chromosome and look where the DNA entered chromosome through double crossover * produce a plasmid that has sequences homologous to a target site in the chromosome* -used to turn genes off by making a frameshift which inactivates that particular gene -put plasmid growing in cells in tissue culture that s in an antibiotic medium > this shuts down native gene because only the cells containing the plasmid are antibiotic resistant this is called a KNOCKOUT -you can do this w eukaryotes in tissue culture but you can't get animals back from it but only in mice - use embryonic stem cells and grow them in tissue culture and put it into an already grown embryo and these stem cells will participate in making a embryo -used for studies for models for human disease like muscular dystrophy and cystic fibrosis
Two types of Gene Libraries
*Genomic DNA libraries -- cloned fragments of DNA isolated from chromosomes; represents all DNA sequences -used for cloning regulatory sequences (promoters), studying genome organization, sequencing of whole genomes, if you want to find introns *cDNA libraries -- cloned DNA copies of mRNA isolated from cells or tissues -used for isolating expressed genes -used to study genes expressed in certain cells and tissues under certain conditions
Analysis of cloned genes - Blotting - Use blotting to see how the gene is put together inside the cell or use blotting to measure expression
*Southern blotting is where DNA is cut with a restriction enzyme, run on a gel, and probed for a specific sequence, used to study structure of a gene or chromosome -used for which clones in a library contain a given DNA sequence *Northern blotting isolates RNA (takes specific mRNA from a certain type of cell/tissue, separated by gel electrophoresis, then transferred to a membrane to be probed) and then is run on a gel, and probed for a specific sequence - to study gene expression ***probes for the presence of mRNA complementary to a cloned gene
Because of genetically engineered foods...
*we've had genetically modified crops approved for human consumption since 1990, improvements in agriculture and yields has plateaued probably -Crop yields have more than quadrupled in the last century (developed countries) -Half of the increase has been due to improvements in agricultural practices, half to plant breeding -In the future genetic engineering is going to play an Increasingly important role
Two competing Groups Sequenced the Human Genome: Celera Inc. (Craig Venter) and International Human Genome Consortium (Francis Collins)
-"Draft" Sequence Published Feb 2001 -"Finished" Sequence 2003 -human genome project was an international project - worked collaboratively - Celera company helped in the late 1990's and sequenced faster and cheaper -these 2 groups were competing against each other, in 2000, clinton got them to negotiate and publish there data together at the same time in the same month ***Celera just wanted to identify as many genes as possible so they dropped out of the project, and human genome consortium finished the job
Genetic and Fossil Evidence Suggests Modern Humans Migrated out of Africa in Two Waves
-70,000 years ago to India, Southern Asia and Australia -50,000 years ago to Europe and Northern Asia -Europe and Asia were already occupied -- Neanderthals
Genetic Modification Of Plants Using Agrobacterium tumefaciens Crown Gall Disease
-Agrobacterium carries a plasmid and transfers a piece of that plasmid into plant cells upon infection (transfer T-DNA) -the T-DNA contains genes that stimulate auxin and cytokinin synthesis in infected plant tissues -Best way of introduction of new DNA uses natural soil bacteria that is a cellular overgrowth - tumor induced plasmid and transfers it into plant chromosome and the piece of DNA that goes into plant cell contains genes that messes up plant hormone production causing the mass of cells -people found that you can take out whatever DNA you want and insert it into the Tumor induced plasmid (Ti) which will become part of the original plant chromosome
Cloning by PCR - Polymerase Chain Reaction
-Allows amplification of specific DNA sequences -Used to clone genes without making a library -Wide range of applications in genetics, molecular biology, evolution, medicine, criminal cases
Retroviral Oncogenes are altered versions of normal cellular genes
-Altered activity or over expression of oncogenes results in cancer (gain of function mutations) -Only one allele of a proto-oncogene needs to be mutated or misexpressed in order to trigger uncontrolled growth SO oncogenes are a dominant cancer phenotype *viral oncogene is an oncogene in a virus
Gene Therapy for ADA/SCID
-Bacterium carrying plasmid with cloned normal human ADA gene and a genetically disabled retrovirus are coupled so the cloned ADA gene is incorporated into the virus -bone marrow cells are removed and cultured from SCID patient, and the engineered retrovirus is added to cell cultures -the retrovirus infects blood cells, transfers ADA gene to cells -genetically modified bone marrow cells are bulked up and returned to patient and see if you get a therapeutic response
Bacillus thuringiensis - BT toxin Insect Resistance
-Bt is a naturally occurring soil bacterium that produces a protein that is toxic to insects -BT has been used as an insecticide for over 100 years, without evidence of environmental problems or human or animal toxicity -Organic farmers use BT dust for insect control *BT causes the digestive tract of insects that ingest it to lyse -effective against caterpillar/mosquitoes/beetles - used in agriculture for over 100 years -Comes in the form of powder which is a dry bacterial culture -Genetically modified plants that makes BT makes its usage easier
Amplification of MYC and MDM2 in Neuroblastoma cells
-Cancer cells have genomic instability -increase in chromosome number -amplification of specific bits of the chromosome -in this picture the 2 cancer genes are amplified as small DNA fragments that remain separate from the chromosomal DNA within the nucleus -MYC is red and MDM2 is green -Normal chromosomes are blue -With MYC, you have pieces of DNA that have gotten off the chromosome and replicated in left -in the right there is amplification on the chromosome increasing the protein product from these cancer causing cells
Carcinogens cause cancer by causing mutations
-Cancer is a genetic disease - what we know is that the association between cancer and mutations is tight ALL OF THE FOLLOWING ARE CARCINOGENS THAT CAUSE CANCER: -Radiation -Polycyclic hydrocarbons -benzopyrene -Nitrosamines -Aflatoxin
Cancer cells exhibit: *loss of growth control - cells continue to divide *altered cell appearance - partially dedifferentiated *ability to move to new site - metastasis
-Cancer is due to accumulation of mutations in somatic cells of genes involved in growth control -Cancer is a diverse set of diseases, many types -general features of all cancers: loss of growth control so cells continue to divide when they shouldn't, there is altered cellular appearance because they're expressing a different set of genes which alters their phenotype so they look different, cells lose their identity and move to different sites in the body, usually they stay in the same specific area (metastasis) We know that cancer is due to the accumulation of genes in growth control MITOSIS
Chapter 16 -- Cancer
-Cancer results from interaction of genes and environment -Today, the most frequent is skin, then lung -if you looked at the same list of cancer frequencies from the1900s, the list would be the same except lung cancer would be number 10 instead of 2
Evidence that Cancer is a Genetic Disease
-Carcinogens are mutagens -Predisposition to cancer is inherited in some families: Mutations in genes encoding DNA repair enzymes result in predisposition to cancer -Specific chromosome rearrangements are seen in some cancers: CML and the "Philadelphia chromosome" -Cancer cells show genomic instability: Translocations, deletions, gene amplifications *Cancer causing chemicals are mutagenic - you can measure how frequently chemicals cause damage to DNA by measuring the rate at which you treat salmonella with the chemical and measure how much mutations you get -predisposition to cancer can be inherited, in particular the enzymes for DNA repair promotes cancer -in some cancers you have very specific chromosome rearrangements which suggests that typical genetic rearrangements are causing cancer -cancer cells show wide scale genomic instability: insertions, deletions, amplifications, translocations - creating abnormal cells with weird genetic expression
Philadelphia Chromosome - Chronic Myelogenous Leukemia
-Caused by a reciprocal translocation between chromosome 9 and 22 - placing BCR and ABL gene next to each other, now cell division regulator ABL is under control of promoter of BCR and then you'll have over production of ABL which kicks off the cancer -presence of this translocation causes the cancer 95% of the time -IT IS SPECIFICALLY THE TRANSLOCATED 22 CHROMOSOME WITH A SMALL PIECE OF CHROMOSOME 9 ON THE BOTTOM OF IT (9 has the ABL gene)
Next Gen Sequencing the quest for the $1000 genome
-Current work focuses on sequencing different individuals to see how we differ and how these differences correlate with health and disease --- We are all 99.9% the same -The search for SNPs - single nucleotide polymorphisms - single base changes in the genome that usually cause disease -Sequencing has be revolutionized: cost came way down, and so this opens new avenues for new things, start looking at what the differences are between humans and how does this relate to disease -Look for snaps that correlate loss of functionality (SNP) - even at this cost it would take 30,000$ to sequence the genome 8 times to be accurate - only need to sequence coding part which you can do for less than 1000$
Southern blotting
-DNA can come from several sources -DNA samples are cut into fragments using 1 or more restriction enzymes, fragments are loaded onto agarose gel and separated by gel electrophoresis -The DNA in the gel is denatured with alkaline treatment to form single stranded fragments -Gel is then overlaid with DNA binding membrane (usually nylon) -To get the DNA fragments to the membrane by placing this on top of a wick (sponge) in buffer solution (gel is directly on top of sponge with membrane on top of it), with paper towels and weight on top of it - this draws buffer up through the gel, transferring the DNA fragments to the membrane -Membrane (filter) is placed in a heat sealed food bag with solution containing single stranded radioactive DNA probe; probe hybridizes with complementary seqeunces -Membrane is washed to removed unbound probe, dried, x-ray film is applied
Human Genome Consortium
-DNA from 8 males was sequenced - 23.1 billion bp sequenced -Human genome size - 3.2 Gb only 2.9 Gb was sequenced: centromeres and telomeres were skipped because they have very little DNA and are very repeated, they are difficult to sequence because of so many repeats so the areas around them are ignored -Gaps - about 5% of the sequence was in gaps between clones, Sequencing accuracy about 99.9% - "draft genome" -"Complete Sequence" 2003: 99.99% accuracy, some 300 gaps still remained -Human genome project doesn't say where the DNA came from - did it 8 fold coverage: didn't sequence females because they're missing the Y -In 2001 when the published they had 95% of the sequence, 5% had gaps, accuracy of 99.9% - this is draft sequence -2003 human genome project is done - they got it to 99.99% accuracy - about 300 gaps remaining, can't have it all but have most of it
Celera
-DNA from five subjects was selected for genomic DNA sequencing: two males and three females -one African-American, one Asian-Chinese, one Hispanic-Mexican, and two Caucasians -27.3 million sequence reads; 14.9 billion bp of sequence -when they published, Celera said they had DNA from 5 different people, different ethnic mix, human genome is only 3 billion base pairs but you have to sequence it multiple times to get an accurate reading with no gaps, sequenced each genome 5 times, which is barely enough to get a draft sequence
Genomic DNA Cut with a Restriction Enzyme Individual DNA Bands are not usually seen Because there are so many different fragments
-DNA has been cut with different restriction enzymes then run on different lanes, and the DNA has been stained w fluorescent dye, the right shows bands containing DNA sequences complementary to probe SHOWING hybridization -No bands on left because when you take DNA and cut w ECOR1 you expect to get a lot of fragments of different lengths, so the bands are in there but theres so many they appear as a smear ****SIZE OF A BAND SHOES THE SIZE OF A RESTRICTION SITE
You can also use Allele-Specific Oligonucleotides for genetic testing to find single nucleotide differences, but that are not limited to just the restriction enzyme cutting sites (like with RFLPs)
-DNA is extracted and the B globin gene is amplified using PCR, this is then denatured and spotted onto strips of DNA binding membrane and then probed with an ASO for either the normal B globin gene or mutant. If hybridized with the normal and they are homozygous dominant (normal allele, not sickle) then they will have the strongest hybridization, if heterozygous then they will have some hybridization, if homozygous recessive they will have no hybridization to the probe *only identical nucleotide sequences can hybridize to ASO -Way to detect single base pair substitution: using hybridization using specific short pieces of DNA -This is for sickle cell -Used to detect single base seqeunces so you can distinguish these 2 molecules binding to the DNA
Process of analyzing DNA with STRs using PCR
-Each primer set is tagged with 1 fluorescent dyes - red, blue, green, yellow -Each primer is designed to amplify DNA fragments from specific loci, so different loci can have the same color -primers are specific for flanking the STR locus and labeled with blue dye -the double stranded DNA s denatured, primers annealed, and each allele (repeat sequence) is amplified by PCR in the presence of dNTPS and Taq DNA polymerase > then separated by cap electro -the sizes of amplified DNA fragments from one color can help differentiate the loci products that have the same color -after amplification, the sizes of amplified fragments are measured by capillary electrophoresis - uses a capillary tube with gel to run a current through it, smaller fragments travel further -a laser detects the fluorescent fragments through the tube -after this the information is translated using computer system finding the size and quantities of each fragment -this DNA profile is then compared to that of other peoples
Genetic Testing for Point Mutations
-Genetic detection of Sickle cell anemia -MstII RFLP -Use this to detect individuals that are heterozygous, homozygous recessive, or homozygous dominant for sickle cell anemia -Use the region where the point mutation can occur in the restriction site as a probe, for the first exon of the gene - if it is normal - you get 2 bands (GAG) mutant is (GTG) *This only happens if the single base change is right in the restrction enzyme site which is unlikely to be the case but it can happen
Transgenic Organisms
-Genetic modification of bacteria, animals, and plants -Introduction of genes from other organisms -clone gene from donor organism into a plasmid -modify gene sequence to permit expression in target organism -introduce plasmid into cells of target organism
Chapter 19: Applications of Gene cloning Recombinant DNA
-Identification and Isolation of Important Genes in Humans and other Organisms -Screening for genetic disorders -DNA fingerprinting -- identification of individuals -Biotechnology -- genetic alteration of animals, plants, and bacteria -Gene Therapy -- correcting genetic disorders
RFLPs allow detection of inheritance of chromosome segments in families that are linked to a disease gene
-Inheritance of RFLPs is Codominant -RFLP allows u to detect the inheritance for genetic variance - imagine you have an RFLP that's inherited with association with an inherited disorder, take DNA from diff people cut with a restriction enzyme and hybridize with a probe > detect diff in where the enzyme cuts bc you'll have slightly different sequences in each -probe needs to be close or linked to gene you're seeing you might have, nearby will work too -Detecting genotype of people with respect to disorder is easy -Use southern blot to see the difference in fragments that are cut using restriction enzymes - DNA fragments are separated with gel electrophoresis, transferred to a nylon membrane, visualized with southern blot hybridization, and probed for mutation region
Gene Sequence Comparisons Help Identify Gene Function: Human NF1 and Yeast Ira Genes
-Ira and NF1 proteins both regulate the Ras protein -More and more genes are cloned and studied from different organisms, and knowing the functions of some of these genes in one organism can help predict the function of these genes in a different organism ***this is a comparison at the amino acid level with a human NF1 gene and the yeast ira gene, the NF1 gene was cloned years ago but no one knew what this protein did, when the yeast ira sequence was cloned, it turned out that it was similar to NF1 and ira gene has a known function: SO we now know about NF1 gene too -the yellow boxes are identical matches between the 2 genes, the lines are chemically similar but not identical -20-30% matching is actually significant, 5% is random similarities, you expect similarities because of a lot of mutations have occured since we evolved from this yeast ***organisms are used as model systems to be informative about the human genome
To explain ^
-Limitations in PCR: not specific in finding a sequence - primers are hard to design, make sure primers don't hit multiple sites in genome, the temperature where you anneal is really important -Make sure primers don't bind to off target sites and if they do, after 1 round of replication, it will bind perfectly to non perfect site -Anneal temperate based on MP of primers, run 3 degrees cooler than what you find it to be -You can amplify something unintended -enzymes can make errors: taq polymerase doesn't have proof reading function, will introduce errors into DNA that you're copying - when it puts in wrong base it stops replicating and reaction terminates, can't copy long pieces of DNA with taq polymerase bc of error rate - to get around this they do a long pcr with an enzyme w proof reading function
The difficulties with human gene therapy primarily have to do with delivery of the DNA to sufficient numbers of target cells in the patient to get a therapeutic response, generally less than 10% of treated cells
-MLV - only infects dividing cells, tends to integrate in transcriptionally active regions - cancer prone -Lentiviruses (HIV) - infect both dividing and nondividing cells, less likely to integrate in transcriptionally active regions -Adenovirus and AAV - DNA genomes, do not integrate into the chromosomes but will infect nondividing and dividing cells -Viruses can trigger immune responses in the patient particularly if the patient has been exposed to the virus before (adenovirus particularly bad in this way)
Microsatellites (STRs/ short tandem repeats) are used for DNA Fingerprinting
-Microsatellites - 2 to 7 base sequences repeated less than 50 times -Different individuals can differ in the number of copies at a given site. -"STRs" - short tandem repeats -known locus STR D8S1179 is made up of a 4-base pair sequence, TCTA, repeated 7-20 times depending on the allele (number of copies of STR)
Cloning the gene for Huntington's Disease -- "Positional Cloning"
-Nancy Wexler cloned the Huntington's gene in 1991 -You look for linkage between inheritance of the disorder and DNA markers that have been mapped to specific sites on chromosomes -5000 related individuals in Venezula who were segregating for HD - including 100 who had the disease -FIRST SUCCESS -Nothing was known about Huntington's disease except that it was inherited - she looked for the disorder and inheritance of RFLPs, map it -connection between the dominant allele for a disease and how that RFLP in the dominant allele, recessive allele is cut differently and is not linked to the disease
Many important genes have been cloned using mapped RFLPs
-Neurofibromatosis -Muscular Dystrophy -Cystic Fibrosis -Huntingtons disease -Proof that the correct gene has been cloned?? -individuals with the disorder will have mutations in the candidate gene -Proof would be that individuals with the disorder have some sequence that people that don't have it don't have
GENETIC SCREENING
-Once a Gene for a disorder has been cloned the cloned gene can be used to identify individuals with the disorder using RFLPs *Detection of b-thalassemia - for the beta globin gene -Deletion or insertion mutations are easiest to detect In this version, there is a deletion that removes exon 3 > detected because there is an enzyme that cuts on either side of beta globin gene, and people with the mutation to cause sickle cell, like here, there band is shorter than with people with the normal allele, aka it'll be closer to the bottom -use pattern matching to detect mutant -All grey means you have 2 normal alleles -This method works well if mutant allele has big insertion or deletion in it, which is the case for this beta knot (mutant allele) -A single base change would have to be right in the restriction site to detect it
Analysis of cloned genes - restriction mapping
-Once you clone a gene you can make a map of it using restriction enzymes -Once you clone a gene you break it up into smaller pieces to take a closer look at it > and that is done with restriction maps -Clone DNA into a plasmid > cut plasmid with different restriction enzymes so you can make a restriction map and see where they are cut within the plasmid -If you get 2 size markers, that means the enzyme cut twice -The size markers indicate how far the cuts are from each other: EcoRI cuts twice at 7 and 3, so the distance between the 2 cuts in 1 direction will add up to 7, and the other add up to 3
To identify genes the computers look for ORFs (open reading frames) and other features of genes
-Once you have the sequence the job is finding the genes, you can use computers algorithms to find sequences consistent w genes, look for open reading frames with no stop codons -Open reading frames have no stop codons aka its probably exons, and identify exons this way, look for promoter sequences/splicing seqeunces -train computers to find this, and find the likely genes, this is annotating the genes, if an annotated gene maps back to an EST then you really have the right gene, verifies the annotation GOAL: FIND ALL THE GENES
Most oncogenes are involved in the hormone signal pathways leading to expression of genes that stimulate cell division
-Oncogenes are genes whose normal function is to promote cell division (under appropriate conditions) -Most oncogenes are genes that code for proteins that code for hormone response -all cell division is regulated by these hormone responses that promote or inhibit cell division -Ras proteins transmit signals from the cell membrane to the nucleus, stimulating the cell to divide in response to growth factors -Ras protein switches off from active (GTP) and inactive form when growth factors aren't binded (GDP) -growth factors bind to the growth factor receptors on the cell membrane, activating Ras -the active GTP bound form of Ras then sends its signals through cascades of protein phosphorylations in the cytoplasm -the end point of these cascades activates activate nuclear transcription factors that stimulate expression of genes whose products start the cell into the cell cycle -once Ras sends its signal to nucleus, it transforms active GTP to GDP, to inactive it -mutations of proto-oncogene Ras into an oncogene will prevent it from turning GTP into GDP, so the cell thinks there is always a growth factor present and always divides **mutations in any of the Ras signaling pathway phosphorylations make them stimulated all the time in the absence of Ras, making them potential Oncogenes
Disadvantages of PCR
-PCR is not necessarily specific for the intended sequence -The primers and annealing temperature determine the specificity of PCR -Short versus Long PCR - PCR is not necessarily accurate -Taq Polymerase lacks 3'-5' exonuclease activity
PCR is a rapid method of DNA cloning > eliminates the use for host cells for cloning
-PCR used to amplify target DNA seqeunces that are initially present in small quantities -double stranded target DNA + DNA polymerase + and 4 DNA bases (A/T/C/G) are added to test tube -some info must be known about target DNA in order to synthesize 2 oligonucleotide primers (single stranded) -these primers bind to complementary DNA after they have been denatured, and are extended by DNA polymerase at the 3' end to make a clone -each cycle doubles the number of DNA molecules in the reaction -Specific DNA polymerase is used bc not all of them can handle the temperature change of annealing and cooling - Taq polymerase -DNA used can come from many samples - even used for forensics like hair samples and dried blood
Virus Resistance: transgenic plants expressing a piece of the viral genome are often resistant to the virus
-Particularly effective with RNA viruses and the mechanism of resistance involves RNA interference ***Papayas and Squash -In the 1990's Hawaiian Papaya production dropped by more than 50% due to Papaya ringspot virus. -Resistant varieties began to be planted in 2000 and the Papaya industry has recovered -this was a success in papayas -RNA interference (RNAi) is a biological process in which RNA molecules inhibit gene expression, typically by causing the destruction of siRNA mRNA molecules. -siRNA comes from the binding of the virus with the RNA and then the dicer cuts it up and the RITS inactivates it -Use genetic modifications to build RNA response to a virus so the plant is ready for the virus before it comes - this is done with RNA interference
DNA Fingerprinting -- Identification of Individuals
-RFLP differences between individuals -VNTRs - Variable Number Tandem Repeats "Minisatellites" short sequences (2-50 bases long) multiple copies at many different sites individuals vary in the number of copies 2-100 copies per site at ~ 40 sites -FE, if you examined 4 different VNTR loci, and each had 20 possible alleles, there would be 4^20 possible genotypes in this profile -someones unique DNA profile can be found for each person if 5-6 loci are analyzed - this is due to the large number of possible VNTRs and alleles -different alleles > from different repeat lengths
Restriction Enzymes
-Restriction endonucleases, cut DNA in a sequence specific manner -Recognition sites are Palindromic sequences: nucleotide sequence reads the same on both strands of the DNA when read in the 5' to 3' direction. -Each restriction enzyme recognizes a specific restriction site and cuts the DNA in a specific way **G/A cut: enzyme does this. Then theres a complementary overhanging ends called sticky ends, and these will be used for reassociation (different pieces from different sources) and they will stick to each other using complementary base pairing. -EcoRI have sticky ends, the vector must also been cut by the same restriction enzyme
New methods of DNA Sequencing are Driving down the cost and time required. The hope is that we will soon be able to sequence individual genomes for $1000
-Revolutions in DNA sequencing - ways to make DNA sequencing cheaper and faster > goal is to bring the cost of sequencing genomes down -Human genome sequencing itself took 10 years and took 3 or 5 billion dollars > now it can be done in a matter of days -You can target specific spots in genome and get a lot of info into medical background
Human Gene Therapy - aims to transfer normal genes into a patients cells
-SCID -- ADA deficiency, SCIDX -The Bubble-boy died of SCID at age 11 Good Candidate Disorders for Gene Therapy Trials? *Simple recessive disorder due to a single gene *Life threatening with little or no other treatment available *Disorder of the circulatory system - SCID - people born w recessive disorders that eliminates their immune system -People with SCID are sensitive to any infection that comes their way -There are 2 different mutations than can cause SCID -This seemed like good candidate for gene therapy in humans - it is a simple recessive disorder for 1 gene (put in correct gene), the disorder is life threatening with little or no other treatment, and this is a disorder of the circulatory system - so you can take cells from the bone marrow and alter them and put them back -anything with the circulatory system should be fixable w gene therapy
Golden Rice - Increased Vitamin A content (carotenoid synthesis genes from daffodil)
-Soybeans and canola with monounsaturated Oleic acid instead of polyunsaturated oils -It is possible to change the nutritional value of things - like in rice. It is genetically modified to increase the expression of vitamin A content
Several Types of "Vectors" are used in cloning, they differ in the size of the "inserts" they can carry
-Standard Plasmids -- inserts < 10 kbp -Bacteriophage Vectors -- inserts 10-50 kbp -BAC Vectors -- inserts 100 kbp, ***All to know- different vectors are used to carry different size pieces of dna, so this makes them used for different purposes. -standard bacterial plasmids can hold about 10,000 bases of dna -Viruses-lambda- can hold a piece of DNA up to about 50,000 base pairs, typical size 25,000. this is a virus where its lysogenic, enters bacterial cell and becomes dormant. So to use bacteriophage as a vector, what has been done is to isolate the pieces at the end and you don't need the genes in the middle. Use these ends to clone DNA into it and the virus will package it. The f plasmid- only part is needed is the part for replication. ***lysogeny - viral DNA is integrated into bacterial chromosome, so every time the chromosome is replicated the viral DNA is replicated and passed down
Exome Sequencing - Personalized DNA Sequencing
-Take DNA from an individual and hybridize it to a gene chip containing sequences that match all human exons (protein coding genes). -After hybridization, recover the exons from the slide and sequence them. -Cost ~ $1,000 (1% of the genome but all the genes) -Hybridize DNA onto a slide and where u have spotted oligonucleotides that correspond to every exon, wash the slide to wash away anything that different hybridize, then wash it so the exons come off, take the exons and sequence it -Used to screen tumors to find an effective treatment
RFLPs Can be Mapped - Mapped RFLPs can be used to map and then clone genetic disorders
-Take a 3 point cross from 2 different people and you have different probes and fragment sizes and you can look to see if there is linkage between different alleles with these RFLPs -anything that says D1 is a RFLP site mapped to chromosome 1 - once they're mapped you can look at inheritance of RFLP and alleles of genes
Cloning DNA yields a "gene library" - collection of clones that contains all the DNA sequences of an organisms genome
-The "library" is a collection of clones 1. Isolated DNA from the organism 2. Cut DNA with Restriction Enzymes 3. Insert DNA fragments into a "Vector" (plasmid) 4. Introduce recombinant DNA into host cell E. coli) 5. Each bacterial colony contains a single recombinant DNA clone *restriction enzyme used for whole genomes is YAC Major challenge: how to identify the specific clone that has the gene you want? Screening Libraries
2009 - successful correction of X-linked adrenoleukodystrophy (ALD) in two young boys using a lentivirus (retrovirus that can infect non-dividing cells)
-Thousands of clinical trials are underway to treat cystic fibrosis hemophilia Parkinsons HIV and more -there is not a cancer problem with this, the problem is DNA delivery
How to make transgenic animals
-Transfection - cloned DNA Introduced into tissue culture cells -Add target DNA to cell culture in pea tree dish -DNA enters nucleus and is expressed but is not integrated into chromosome -1 colony forms/10^6 cells treated with 40 pg gene DNA *DNA must integrate into a chromosome to be maintained and stably expressed *The site of DNA integration is essentially random *DNA integration may cause a mutation *****Site where new DNA is integrated occurs via nonhomologous DNA repair mechanism, but it puts this piece of DNA anywhere, which is a problem that was only solved in mice
Making transgenic mice
-Transgenic Mice by injection of DNA into a fertilized egg -then the fertilized eggs are placed in the oviduct of a receptive (previously pregnant) female -RECOMBINANT DNA CAN BE PUT INTO A SPECIFIC SPOT IN MICE
*****Genetic corrections of cells in the germ line are ruled out because they would be heritable putting future generations at risk.
-Treatment is targeted to specific organs in the patient -Disorders of the circulatory system are most amenable because bone marrow stem cells can be removed and grown in tissue culture **in the circulatory system you can take cells out, make genetic modifications to them in a dish, and then put the cells back > the biggest problem is to get DNA into the target organism in big enough amounts to fix the problem - delivery mechanism is tailored to the organism
Predictions based on annotation and analysis of protein functional domains and motifs: genes in the human genome that have been assigned on the basis of similarity to proteins of known function (in other organisms) (slide 11 set 31)
-Use computers to predict what the unknown genes do using blast to compare predicted (unknown) gene to everything else that has ever been cloned that has info for what that gene does - if 2 genes are similar according to blast, they have proteins with similar function -Doesn't tell you which genes it codes for only tells us that it codes for a specific transcription factor, not which ones -molecular function unknown: 40% unknown, we don't know what is expressed and coded for by these genes
DNA is cut with a restriction enzyme that cuts outside of the VNTR
-VNTRs have codominant inheritance - so you'll see the bands for each allele at each loci they're testing -take a piece of DNA, cut it with a restriction enzyme that cleaves on either side of the VNTR repeat region -the digested DNA is separated by gel electrophoresis and then run on a southern blot -briefly, separated DNA is transferred from the gel to a membrane and hybridized with a radioactive probe that recognizes sequences within the VNTR region -expose membrane to x-ray film, the pattern of bands is measured, with larger VNTR repeat alleles remaining at the top of the gel, and smaller VNTRs which migrate more rapidly, being closer to the bottom ***THE LENGTH OF THE VNTR REPEATS REFLECTS A DIFFERENT ALLELE
Retrovirus Life Cycle
-Viral RNA enters host cell -DNA copy of Virus made -Viral DNA integrates into host chromosome -Viral RNA made by transcription -Release of new viral particles (PROTEINS)
Plants and Mammals are also used as "bioreactors" for production of human proteins because bacteria do not glycosylate proteins
-bioreactors - living factories - that will continuously make the product containing the desired therapeutic protein that can be isolated in a noninvasive way -regardless of the host, therapeutic proteins may then be purified from host cells
Overproduction of the Gene for Growth Hormone in Fish
-contain transgenic growth hormone genes -transgenic atlantic salmon have copies of chinook salmon growth hormone next to a constitutive expressed (unregulated, constantly active) promoter -no adverse health effects from added gene
Expression of b-globin genes during development
-different members of the beta gene family have slightly different functions, and are active in their own time in development -People with sickle cell > treat it to improve expression of delta because it will make a normal beta chain and increase the good beta in people with sick cell disease
Retroviruses can acquire host genes
-first you have a nonacute retrovirus - the genome of the virus can infect the cell but not transform it -the retrovirus can then pick up a copy of the host proto-oncogene during its infection of the cell and integrate it into its genome -the cellular proto-oncogene may be mutated during the process of transfer into the virus, or it may be expressed at abnormal levels because it is now under the control of viral promoters -these new retroviruses can infect and transform normal cells into tumor cells, they are ACUTE TRANSFORMING RETROVIRUSES *the transfer of gene from host cell into the viral genome allows the virus to infect and transform cells MAKING IT AN ONCOGENIC RETROVIRUS -An acute retrovirus has an additional part - these are the genes that cause cancer, somehow the virus picked up a normal gene while infecting the host cell, which mutates normal gene and causes cancer -This can happen because the virus planks down into chromosome and gene is right next door, the neighboring piece of DNA can end up in the retrovirus during its replication
GWAS - Genome Wide Association Studies -The search for genes responsible for complex traits using SNPs FOR EXAMPLE - Type II diabetes
-genomes of thousands of unrelated people with a particular disease are analyzed, usually using microarray analysis -results are compared with people without the disease to identify genetic variations that may increase risk of getting the disease -utilize large scale use of SNP microarrays that probe for 500,000 SNPs to evaluate results from diff people -by looking at SNP changes that occur in people with the disease, scientists can calculate the disease risk associated with each genetic variation using statistics
Transgenic plants are made using the soil bacterium Agrobacterium tumefaciens -bacterial EPSP synthase gene provides resistance to the broad spectrum herbicide Glyphosate
-herbicides kill plants by inhibiting the action of the enzyme EPSP sythase -to make herbicide resistant plants you ... -take the EPSP synthase gene from bacteria and fuse it to the right promoter and poly A signals CREATING a fusion gene -introduce fusion gene into TI plasmid that doesn't have tumor genes and then this is transformed into host plant cells in tissue culture -Plants expressing the EPSP synthase gene are grown from the tissue culture - you know they're expressing it if they are survive on glyphosphate medium *cells that acquire the EPSP synthase gene are able to synthesize large quantities of it, making them resistant to herbicide glyphosate -use a promoter that comes from a virus to obtain a high level of expression in all cells throughout the plant, use specific promoters if you want it to be specific placement like in leaves or stems, etc
-Genes are separated by regions of spacer DNA -Genes are on both strands, DNA on both strands can code -Genes on different strands can overlap -If there is the same gene running in the opposite direction on the opposite strand > it codes for a different protein
-human have fewer overlapping genes than simpler eukaryotes
Limitations of PCR
-info about DNA is needed to make primers -minor contamination of DNA can cause problems for accuracy (ex: skin cells from researcher contaminate samples from crime scene) -Usually, DNA poly in PCR only extends primers for short distances, not to the end of the strand, SO PCR is usually used to amplify relatively short pieces of DNA
Transgenic Animals -- mice, sheep, pigs, etc *Microinjection of DNA into a Fertilized Egg *Egg is implanted into uterus
-isolate newly fertilized eggs in host female animal -inject purified cloned DNA containing a vector and the transgene of interest into nucleus of egg -only a small amount of the transgenic eggs will contain the transgenic DNA into the egg cell genome -injected eggs are then placed into the oviduct of a animal previously impregnated by a male -once baby is born it is screened for the trans gene via PCR -the integrated DNA must be present in germ (gamete) cells so it is passed to all offspring -most F1 aren't homozygous for the transgene so the F1 siblings must mate to make homozygous transgenic animals
Eukaryotic genomes
-linear, genes less densely packed, -monocistronic with introns in the genes -amount of repeat DNA and intron size increase with evolutionary complexity -Amount of noncoding spacer DNA and the number and size of introns increases with evolutionary complexity in eukaryotes -the average gene has 12 introns in it -Duchenne Muscular Dystrophy gene -- 2 Mb, >70 introns! ***biggest gene
Gene Chips - Microarrays
-oligonucleotides specific for different genes are spotted onto a slide - 64,000 genes on an area the size of a microscope slide coverslip -used for looking at gene expression patterns in genetic diseases -may use probes for only a few specific genes thought to be expressed in different cell types OR may contain probes for each gene in the genome -On this chip, pieces of DNA that will hybridize to specific probes are used to look at different gene expression in different tissues -Take normal cells and tumor cells, isolate mRNA in both, change them into cDNA, and treat them with fluorescent dyes, amount of each cDNA is proportional to amount of mRNA - mix fluorescent cDNAs and wash and let them dry and hybridize and they'll find the oligonucleotide they correspond to - laser and measure the amount of fluorescence in each, which is the output *****the probes are already on the chip/microarray for certain genes you're looking at, so when you add the fluorescent cDNAs, the cDNAs hybridize to the corresponding field gene that encodes their mRNA -yellow means they are expressed, or present in the same probe gene -you have a gene as the probe, whatever hybridizes to it (the color) tells you if the mRNA is expressed in normal or cancer cells -More red > tumor -More green > normal -Yellow has equal amount of mRNA in normal and cancer cells - level of expression of the same gene in both normal and cancer cells -Used to see which genes have been turned on or off in tumor - red is the gene turned on in cancer cells -If an mRNA is present only in normal cells, the probe representing the gene encoding that mRNA will appear as a green dot because only the "green" cDNA have hybridized to it
In 2009 the FDA Approved transgenic Goats for the production Of anticoagulant
-put human anticoagulant gene next to promoter for beta caesin, a common protein in milk, they were able to target anticoagulant expression in the mammary gland, so this is highly expressed in the milk, and then isolate it from the milk
Some genes in eukaryotes are present in multiple copies - like for rRNA and tRNA -slide set 31 slide 19
-rRNAs 500-5000 copies in tandem arrays -tRNAs 2 to 20 copies in clusters -coding genes themselves are copies -We need multiple gene copies because there is no amplification process for making rRNA/tRNA SO to have enough rRNA for ribosomes we need to have multiple copies of the gene -normal genes can get amplified many times for more multiplication
PCR (powerpoint notes)
-repeated rounds of steps -target DNA and excess of primers and DNA polymerase and the 4 bases, heat up to denature, and then cool the mixture to temp so the primers will just manage to base pair w target and polymerase will start to copy, then heat back up and put it back through the cycle again -doubling of target sequence
Gene Families Evolve by Gene Duplication through Unequal Crossing Over Followed by Sequence Drift
-repeated sequences in the genome - transposable elements - increase the frequency of recombination errors like this -Gene families come from unequal crossing over, if chromosomes are misaligned and crossing over occurs, there will be a duplication on one chromosome and deletion on the other -Duplicating a gene doesn't harm anything so you retain the original gene with a second copy that can take on new function - important in evolution of new genes, let evolution shuffle the parts - this is where transposable elements become important because there presence in the genome creates the errors in crossing over leading to new families - when they align w repeat seqeunces it confuses meiosis mechanisms and forms misalignments when crossing over occurs SO VALUABLE FOR EVOLUTION OF THE GENOME
SNPs - single nucleotide polymorphisms (single nucleotide changes)
-resequencing of the human genome in different individuals has identified 10 million SNPs -SNPs can be used to identify gene variants associated with complex disorders (polygenic traits). -A polygenic trait is one whose phenotype is influenced by more than one gene. Traits that display a continuous distribution, such as height or skin color, are polygenic. -Use these as genetic markers and use these in screening too -May be associated or linked to alleles that cause different disorders -Used to find genes that are associated with complex disorders or polygenic traits, each gene having a tiny effect on increasing the disorder
Bacterial genomes
-small -circular -very little repeated DNA -closely packed protein coding genes -lacks introns -polycistronic genes common but do have monocistronic
RFLPs - only used when there is a mutation in the restriction site, this is the only way you'll see a difference if you run 2 different peoples DNA on a gel
-so you have a cloned DNA segment with an RFLP -if there is a mutant allele, in the presence of a restriction enzyme, it will not be able to cut it, so there will be a large band -for a normal allele there will be 2 smaller bands -so if someone is heterozygous, there is 1 large band for the mutant allele and 2 small bands for the normal -someone with sickle cell (that is homo recessive) will have just 1 large band for both mutant alleles -and homozygous dominant will be 2 smaller bands and thats it
Oligonucleotide synthesis
-ssDNA sequences (up to 40 - 50 bases) are easily synthesized. -Synthetic reactions: add one base at a time and you can make things up for a specific sequence. -short, single stranded fragments of DNA that can be used as a DNA probe for alleles that differ by even a single nucleotide, will hybridize only to complementary base sequence, not even to a sequence off by 1 base
Process of PCR
-start with double stranded DNA to be cloned in test tube with primer (present in excess), DNA polymerase, 4 DNA bases (A/G/C/T) and -first you denature the DNA (heat it up 92-95 deg) -Then the primers anneal to the single stranded DNA (when it is cooled down 45-65 deg) -the primers are extended by DNA polymerase (Taq) at the 3' end, the temperature is raised a little 65-75 deg -That is the end of the first cycle - number of DNA molecules is doubled
Northern Blot of Human RNA from Different Organs: Probed with cloned FMR1 gene (mutations in this gene result in mental retardation)
-take RNA from different cells, run it out on agarose gel, and transfer to a membrane paper, and run it on single stranded DNA probe derived from a cloned copy of a gene -In this case, we are probing for the fmr1 gene (fragile x) RNA from different tissues here and this northern blot tells us that mRNA that corresponds to fmr1 is 4.4 bases long, and that there is more in the brain, placentena, etc and very little in the liver -In the heart the message is small > this may be due to alternate splicing of the mRNA *****Bigger fatter band > more rna that corresponds
Screening a gene library by DNA hybridization using a cloned gene as the "probe"
-the probe is a DNA or RNA sequence that is complementary to the target gene/sequence to be identified in a library 1. colonies of the library are overlaid with a DNA binding membrane such as nylon > this transfers the pattern of bacterial colonies from the plate to the membrane 2. Colonies are transferred to membrane, then lysed and denatured with NaOH 3. Membrane is placed in a heat sealed bag with a solution containing the labeled probe; the probe hybridizes with complementary DNA from colonies 4. Membrane is removed from bag and rinsed to remove excess probe then dried; Xray film is placed over the membrane > hybridization of a clone to colony shows up by a spot on Xray film 5. Colonies containing the inserted probe are identified from the orientation of the spots on the original plate, and removed 6. cells that hybridized with the probe are transferred to a medium for growth and further analysis
How to use a retro virus for gene therapy
-use retrovirus to deliver new gene -the bacterium containing the plasmid with the cloned normal gene > the target gene is inserted into a genetically disabled retrovirus -the retrovirus infects target cells that you want to insert your target gene into and transfers the target gene into it -the infected cells are then cultured to ensure the target gene is active -and then re-implanted back into person -useful because they insert DNA into chromosome
Gene Families - many protein coding genes in eukaryotes are present in multiple copies - similar but not identical (as with alpha and beta)
-we also have multiple copies of protein coding genes, which tells us how genome functions -A gene family is a group of genes that code for similar but not identical proteins EX hemoglobin -We have alpha and beta gene families for hemoglobin, on different chromosomes, there are 5 copies of alpha that are similar but not identical -Chromosome 16 - alpha gene family -Chromosome 11 - beta gene family -On chromosome 11 (beta) - there are 6 copies of that gene all within 50,000 bp, the basic structure of all the genes are very similar, and the length for all beta chains are the same but have amino acid differences between them -Alpha and beta are so incredibly similar -look for surrounding genes to see if it is part of a gene family, lots of pseudo genes ***pseudogenes -- nonfunctional copies of coding genes - found in both alpha and beta -the number of copies includes the pseudo genes too
In mink, brown fur color is dominant to silver-blue fur color. If a homozygous brown mink is mated with a silver-blue mink and 8 offspring are produced. how many would be expected to be silver-blue?
0
Steps in Cloning a Gene
1. Isolated DNA from the organism 2. Cut DNA with Restriction Enzymes 3. Insert DNA fragments into a "Vector" (plasmid) 4. Introduce recombinant DNA (in the form of the plasmid) into host cell E. coli 5. Grow the host and reisolate recombinant DNA clones **to distinguish host cells that have taken up vectors, the vector should carry a slectable marker gene - usually an antiobiotic gene or an enzyme absent from host cell, because then will be obvious when plated on this antiobotic and show they are there
Some Strategies for Screening Libraries
1. Probing using an already cloned gene 2. Isolate the protein, sequence it and make a synthetic oligonucleotide 3. Expression Libraries - antibody detection used
What is catalyzed by a ribozyme
1. cleavage fo precurose RNAs to tRNAs 2. removal of introns during trasn-esterification rxns involved in centain mRNA splicing 3. formation of peptide bonds during protein synthesis
"Shotgun Cloning" approach used by Celera
1. genome cut into small pieces with Restriction Enzymes and cloned 2. clones are sequenced and contigs identified by overlapping end sequences 3. chromosome sequence assembled -Use multiple restriction enzymes, make multiple libraries, and sequence the ends of the pieces, and using this to identify which overlap each other and identify contigs this way instead of physically mapping it, "if 2 pieces have the same end they probably came from same chromosome" -Identified contigs by sequencing the ends then they sequenced the whole pieces and used the computer to put together final chromosome assembly **Computers assemble the sequence of individual contigs into one continuous sequence based on where they overlap/ align (meaning they have the same dna sequence there)
What fraction of this cross will be recessive for both traits?
1/16
Listed below are procedures involved in the production of a transgenic organism. From the choices provided, select the sequence that represents the proper order of events. 1. Recombinant DNA is transferred into a bacterial cell. 2. A specific gene is identified in a DNA sequence. 3. The DNA fragment is recombined into a vector. 4. The DNA fragment to be inserted is isolated.
2, 4, 3, 1
One mechansim used by DNA polymerase to ensure that the inserted base is correct is what? It allows polymerase to backtrackand remove the nuclotide that was most recently incoroporated in to the primer strand
3 to 5 exonucleasue
The effort to completely map and sequence the human genome will likely result in knowing the sequence of the approximately ___ genes on the 46 human chromosomes.
35 000 to 40 000
The effort to completely map and sequence the human genome will likely result in knowing the sequence of the approximately____genes on the 46 chromosomes
35,000- 40,000
A couple has two children, both of whom are boys. What is the chance that the parents' next child will be a boy?
50%
When using DNA fingerprinting to determine the paternity of a child, how many of the child's 13 str loci should match the father's?
6 or 7 -probably because half of your DNA is from dad, and half from mom, so half should match (at each loci, you have an allele from mom, and an allele from dad)
Plants are being "engineered" to be more resistant to insects or viruses, to be herbicide tolerant
75% of soybeans and cotton in the US are herbicide resistant
In chickens, rose comb (R) is dominant to single comb (r). A homozygous rose-combed rooster is mated with a single-combed hen. All of the chicks in the F1 generation were kept together as a group for several years. They were allowed to mate only within their own group. What is the expected phenotype of the F2 chicks?
75% rose comb and 25% single comb
A female guinea pig homozygous dominant for black fur color is mated with a male homozygous for white fur color. In a litter of eight offspring, there would probably be ___.
8 black guinea pigs
Suppose 10 000 units of energy are available at the level of the grasses. What is the total number of energy units lost by the time the third energy level is reached?
9990 units
In Figure 10-8, what gametes will result if there is only a single crossover?
A
In homologous genetic recombination, RecA protein is involved in: A) Strand invasion to form Holliday intermediates. B) introduction of negative supercoils into the recombination products. C) nicking the two duplex DNA molecules to initiate the reaction. D) pairing a DNA strand from one duplex DNA molecule with sequences in another duplex, regardless of complementarity. E) resolution of the Holliday intermediate.
A
Which of the cells depicted in the line graph in Figure 8-8 are most likely cancerous?
A
Which of the following statements about E. coli RNA polymerase is false? A) Core enzyme selectively binds promoter regions, but cannot initiate synthesis without a sigma factor. B) RNA polymerase holoenzyme has several subunits. C) RNA produced by this enzyme will be completely complementary to the DNA template. D) The enzyme adds nucleotides to the 3' end of the growing RNA chain. E) The enzyme cannot synthesize RNA in the absence of DNA.
A
Which structure shown in Figure 11-3 would attract a free cytosine nucleotide?
A
Retrovirus explained and their way of replacing their DNA with the therapeutic DNA
A retrovirus is any virus belonging to the viral family Retroviridae. All The genetic material in retroviruses is in the form of RNA molecules, while the genetic material of their hosts is in the form of DNA. When a retrovirus infects a host cell, it will introduce its RNA together with some enzymes into the cell. This RNA molecule from the retrovirus must produce a DNA copy from its RNA molecule before it can be considered part of the genetic material of the host cell. Retrovirus genomes commonly contain these three open reading frames that encode for proteins that can be found in the mature virus. Group-specific antigen (gag) codes for core and structural proteins of the virus, polymerase (pol) codes for reverse transcriptase, protease and integrase, and envelope (env) codes for the retroviral coat proteins (see figure 1). The process of producing a DNA copy from an RNA molecule is termed reverse transcription. It is carried out by one of the enzymes carried in the virus, called reverse transcriptase. After this DNA copy is produced and is free in the nucleus of the host cell, it must be incorporated into the genome of the host cell. That is, it must be inserted into the large DNA molecules in the cell (the chromosomes). This process is done by another enzyme carried in the virus called integrase (see figure 2). Now that the genetic material of the virus is incorporated and has become part of the genetic material of the host cell, we can say that the host cell is now modified to contain a new gene. If this host cell divides later, its descendants will all contain the new genes. Sometimes the genes of the retrovirus do not express their information immediately. Retroviral vectors are created by removal op the retroviral gag, pol, and env genes. These are replaced by the therapeutic gene. In order to produce vector particles a packaging cell is essential. Packaging cell lines provide all the viral proteins required for capsid production and the virion maturation of the vector. These packaging cell lines have been made so that they contain the gag, pol and env genes. Early packaging cell lines contained replication competent retroviral genomes and a single recombination event between this genome and the retroviral DNA vector could result in the production of a wild type virus. Following insertion of the desired gene into in the retroviral DNA vector, and maintainance of the proper packaging cell line, it is now a simple matter to prepare retroviral vectors (see figure 3). One of the problems of gene therapy using retroviruses is that the integrase enzyme can insert the genetic material of the virus in any arbitrary position in the genome of the host. If genetic material happens to be inserted in the middle of one of the original genes of the host cell, this gene will be disrupted (insertional mutagenesis). If the gene happens to be one regulating cell division, uncontrolled cell division (i.e., cancer) can occur. This problem has recently begun to be addressed by utilizing zinc finger nucleases or by including certain sequences such as the beta-globin locus control region to direct the site of integration to specific chromosomal sites.
DNA fingerprinting
A test to identify and evaluate the genetic information-called DNA in a person's cells
5. Which of the following statements about tRNA molecules is false? A) A, C, G, and U are the only bases present in the molecule. B) Although composed of a single strand of RNA, each molecule contains several short, double-helical regions. C) Any given tRNA will accept only one specific amino acid. D) The amino acid attachment is always to an A nucleotide at the 3' end of the molecule. E) There is at least one tRNA for each of the 20 amino acids.
A) A, C, G, and U are the only bases present in the molecule.
9. Formation of the ribosomal initiation complex for bacterial protein synthesis does not require: A) EF-Tu. B) formylmethionyl tRNAfMet. C) GTP. D) initiation factor 2 (IF-2). E) mRNA.
A) EF-Tu.
3. Which of the following is not one of the five steps of protein synthesis? A) Termination and ribosome degradation B) Elongation of the peptide chain C) Initiation of peptide synthesis D) Activation of the amino acids E) Protein folding and posttranslational processing
A) Termination and ribosome degradation
18. The first step in two-dimensional gel electrophoresis generates a series of protein bands by isoelectric focusing. In a second step, a strip of this gel is turned 90 degrees, placed on another gel containing SDS, and electric current is again applied. In this second step: A) proteins with similar isoelectric points become further separated according to their molecular weights. B) the individual bands become stained so that the isoelectric focus pattern can be visualized. C) the individual bands become visualized by interacting with protein-specific antibodies in the second gel. D) the individual bands undergo a second, more intense isoelectric focusing. E) the proteins in the bands separate more completely because the second electric current is in the opposite polarity to the first current.
A) proteins with similar isoelectric points become further separated according to their molecular weights.
A virus isolated from monkeys contains a circular double strand of DNA. The virus, called Simian Virus 40, interests scientists because it causes cancer in laboratory animals. Using a restriction enzyme, the strand is separated into six unequal segments, as shown in Figure 13-2. A scientist hypothesizes that the segment of DNA causing cancer can contain no fewer than 600 base pairs. Using Figure 13-2, decide which segments of the virus have the highest chance of containing the segment of interest. Identify in DESCENDING order, from the HIGHEST change to the LOWEST.
A, B, C
DNA Markers for Genetic Screening -- RFLPs Restriction Fragment Length Polymorphisms
Base changes or small insertions and deletions that vary between individuals can create or eliminate restriction enzyme sites. (as seen in sickle cell anemia testing)
Making transgenic humans is considered unethical because:
ANSWER: we cannot control where the recombinant DNA will go when it integrates into the chromosomes -You can't control where DNA goes when you add it in, so you create the possibility of mutations in genes that are important - not unethical to make genetic modifications to certain cells, BUT if you make bad mutations in transgenic organisms it can be passed on to the next generation WHICH IS unethical
mRNA leaves the nuclus and enter the cytoplasm. These data supported th yp that
mRNA serves as the template for protein synthesis
The pairing of ___ in DNA is the key feature that allows DNA to be copied.
nitrogen bases
What must be on either end of any genetic material that is inserted in the cleaved DNA in fig 13-5
AATT
What must be on either end of any genetic material that is inserted into the cleaved DNA in Figure 13-5?
AATT
Which of the following disorders would be the easiest to treat by gene therapy and which would be the hardest? 1. Muscular dystrophy 2. Cystic fibrosis 3. Hemophilia
ANSWER: Easiest 3, Hardest 1 easiest to target the circulatory system that is why hemophilia is the answer -harder to target muscles
Cancer is infectious, can be transmitted from person to person
ANSWER: disagree -cancer is not infectious and cannot be transmitted from person to person - one type of cancer that is transmissible is wiping out the Tasmanian devil - if you get cancer cells from someone else your immune system will kill it off, if you have a defective immune system it may be transmissible
Most cancers are caused by viruses
ANSWER: disagree there are some cancers that are caused by viruses, some viruses directly cause cancer, but most cancer doesn't have anything to do with virus
All cancer cells grow faster than normal cells
ANSWER: disagree there are some cancers that are fast growing and some that are slow growing, very diverse, prostate cancer is slow growing
If you have a genetic test to see if you are a carrier of cystic fibrosis and the test comes back negative does this mean your children cannot get the disease?
ANSWER: maybe, maybe not -Difficulty in genetic screening can only detect the known alleles, and we don't know all the alleles that cause the disorder with cystic fibrosis - we know one allele and that accounts for 80% of the people who have it, the other 20% of people who get it don't know where they got it from -Getting back a negative test doesn't mean you're not gonna get it, but a lesser chance, but that u may have a rare allele that can cause it that they didn't test for -If it comes back positive - then you will get it, you have the common allele that causes it
Enzymes called _____________ couple each amino acid to its appropriate tRNA molecule to create an _____________ molecule.
Aminoacyl-tRNA synthetases; Aminoacyl-tRNA
1. The difference between thymine and uracil is: A) one methylene group on the pyrimidine ring. B) one methyl group on the pyrimidine ring. C) one hydroxyl group on the ribose ring. D) one amine group on the pyrimidine ring. E) one methyl group on the purine ring.
B
If the segments in fig 13-6 are mixed with several restriction enzymes, which will not be cleaved
B
Processing of a primary mRNA transcript in a eukaryotic cell does not normally involve: A) attachment of a long poly(A) sequence at the 3' end. B) conversion of normal bases to modified bases, such as inosine and pseudouridine. C) excision of intervening sequences (introns). D) joining of exons. E) methylation of one or more guanine nucleotides at the 5' end.
B
The excision (splicing) of many group I introns requires, in addition to the primary transcript RNA: A) a cytosine nucleoside or nucleotide and a protein enzyme. B) a guanine nucleoside or nucleotide (only). C) a protein enzyme only. D) a small nuclear RNA and a protein enzyme. E) ATP, NAD, and a protein enzyme
B
The role of the Dam methylase is to: A) add a methyl group to uracil, converting it to thymine. B) modify the template strand for recognition by repair systems. C) remove a methyl group from thymine. D) remove a mismatched nucleotide from the template strand. E) replace a mismatched nucleotide with the correct one.
B
Which segment in Figure 13-6 is not a palidrome?
B
Which segment in fig 13-6 is not a palidrome
B
Which segment in fig 13-6 will attach to genetic material with the sequence TCGA
B
Which structure shown in Figure 11-3 is a pyrimidine?
B
10. When a DNA molecule is described as replicating bidirectionally, that means that it has two: A) chains. B) independently replicating segment. C) origins. D) replication forks. E) termination points
D
8. Which of the following is/are true for protein synthesis in eukaryotes? A) All proteins are initially synthesized with methionine at their C-terminus. B) All proteins are initially synthesized with methionine at their N-terminus. C) All proteins are initially synthesized with tryptophan at their C-terminus. D) All proteins are initially synthesized with a multiple of 3 amino acids in their sequence. E) None of the above
B) All proteins are initially synthesized with methionine at their N-terminus.
6. Which of the following is not required for initiation of DNA replication in E. coli? A) DnaB (helicase) B) DnaG (primase) C) Dam methylase D) DNA ligase E) DnaA (a AAA+ ATPase)
D
The alleles present for a trait are the same
homozygous
The diagram in Figure 10-2 shoes a diploid cell with two homologous pairs of chromosomes. Due to independent assortment, the possible allelic combinations that could be found in gametes produced by the meiotic division of this cell are ___.
BD, bD, Bd, and bd
Different Restriction Enzymes cut different DNA sequences
Bacteria that make a restriction enzyme also make a corresponding methylase. This corresponding methylase stops the bacteria from cutting its own DNA at sites that the restriction enzyme would otherwise recognize in the bacteria.
The gamete that contains genes contributed only by the mother is ___.
an egg
There are some inherited predispositions to cancer
Breast Cancer - genes affected are BRCA1, BRCA2 - recombination repair Hereditary nonpolyposis colon cancer - due to mismatch repair XP (xeroderma pigmentosa) - nucleotide excision repair -if you have the mutated gene you are at a greater risk for developing that cancer -Many of these genes affected are involved in DNA repair, causing the mutations and cancer WHICH ties mutation to cancer
9. The Meselson-Stahl experiment established that: A) DNA polymerase has a crucial role in DNA synthesis. B) DNA synthesis in E. coli proceeds by a conservative mechanism. C) DNA synthesis in E. coli proceeds by a semiconservative mechanism. D) DNA synthesis requires dATP, dCTP, dGTP, and dTTP. E) newly synthesized DNA in E. coli has a different base composition than the preexisting DNA.
C
In Figure 10-8, what gametes will result in each chromatid crossed with a nonsister chromatid?
C
In base-excision repair, the first enzyme to act is: A) AP endonuclease. B) Dam methylase. C) DNA glycosylase. D) DNA ligase. E) DNA polymerase.
C
The 5'-terminal cap structure of eukaryotic mRNAs is a(n): A) 7-methylcytosine joined to the mRNA via a 2',3'-cyclic linkage. B) 7-methylguanosine joined to the mRNA via a 5' - 3' diphosphate linkage. C) 7-methylguanosine joined to the mRNA via a 5' - 5' triphosphate linkage. D) N6 -methyladenosine joined to the mRNA via a 5' - 5' phosphodiester bond. E) O6 -methylguanosine joined to the mRNA via a 5' - 5' triphosphate linkage.
C
The PCR reaction mixture does not include: A) all four deoxynucleoside triphosphates. B) DNA containing the sequence to be amplified. C) DNA ligase. D) heat-stable DNA polymerase. E) oligonucleotide primer(s)
C
The repair of cyclobutane pyrimidine dimers by bacterial DNA photolyase involves the cofactor: A) coenzyme A. B) coenzyme Q. C) FADH- D) pyridoxal phosphate (PLP). E) thiamine pyrophosphate (TPP).
C
Which of the following is false about transposition of DNA? A) The diversity of immunoglobins is in part due to DNA recombination by transposition. B) Transposition occurs in both prokaryotes and eukaryotes. C) Enzymes are not required for transposition. D) The first step of transposition can be single- or double-stranded DNA cleavage. E) Transposition can lead to simple movement of a DNA region or duplication of that region in a new location.
C
Listed below are procedure involved in the production of a transgenic organism. From the choices provided, select the sequence that represents the proper order of events.
C - 2,4,3,1
7. At replication forks in E. coli: A) DNA helicases make endonucleolytic cuts in DNA. B) DNA primers are degraded by exonucleases. C) DNA topoisomerases make endonucleolytic cuts in DNA. D) RNA primers are removed by primase. E) RNA primers are synthesized by primase.
E
A branched ("lariat") structure is formed during: A) attachment of a 5' cap to mRNA. B) attachment of poly(A) tails to mRNA. C) processing of preribosomal RNA. D) splicing of all classes of introns. E) splicing of group II introns.
E
14. Which of the following is correct with respect to the amino acid composition of proteins? A) Larger proteins have a more uniform distribution of amino acids than smaller proteins. B) Proteins contain at least one each of the 20 different standard amino acids. C) Proteins with different functions usually differ significantly in their amino acid composition. D) Proteins with the same molecular weight have the same amino acid composition. E) The average molecular weight of an amino acid in a protein increases with the size of the protein.
C) Proteins with different functions usually differ significantly in their amino acid composition.
When bacterial DNA replication introduces a mismatch in a double-stranded DNA, the methyl-directed repair system: A) cannot distinguish the template strand from the newly replicated strand. B) changes both the template strand and the newly replicated strand. C) corrects the DNA strand that is methylated. D) corrects the mismatch by changing the newly replicated strand. E) corrects the mismatch by changing the template strand.
D
Which of the following is not needed in pyrosequencing of DNA? A) dNTPs B) Sulfurylase C) Luciferase D) ddNTPs E) Apyrase
D
Which structure shown in Figure 11-3 does not contain a nitrogenous base?
D
8. In contrast to bacteria, eukaryotic chromosomes need multiple DNA replication origins because: A) eukaryotic chromosomes cannot usually replicate bidirectionally. B) eukaryotic genomes are not usually circular, like the bacterial chromosome is. C) the processivity of the eukaryotic DNA polymerase is much less than the bacterial enzyme. D) their replication rate is much slower, and it would take too long with only a single origin per chromosome. E) they have a variety of DNA polymerases for different purposes, and need a corresponding variety of replication origins.
D
In homologous recombination in E. coli, the protein that moves along a double-stranded DNA, unwinding the strands ahead of it and degrading them, is:
D
The ABC excinuclease is essential in: A) base-excision repair. B) methyl-directed repair. C) mismatch repair. D) nucleotide-excision repair. E) SOS repair.
D
Parasitism, commensalism, and mutualism are examples of ___.
symbiosis
A white mouse whose parents are both white produces only brown offspring when mated with a brown mouse. The white mouse is most probably ___.
homozygous recessive
1. The Ames test is used to: A) detect bacterial viruses. B) determine the rate of DNA replication. C) examine the potency of antibiotics. D) measure the mutagenic effects of various chemical compounds. E) quantify the damaging effects of UV light on DNA molecules.
D
An alternative repair system by error-prone translesion DNA synthesis can result in a high mutation rate, because: A) alternative modified nucleotides can be incorporated more readily. B) interference from the RecA and SSB proteins hinders the normal replication accuracy. C) replication proceeds much faster than normal, resulting in many more mistakes. D) the DNA polymerases involved cannot facilitate base-pairing as well as DNA polymerase III. E) the DNA polymerases involved lack exonuclease proofreading activities.
E
Splicing of introns in nuclear mRNA primary transcripts requires: A) a guanine nucleoside or nucleotide. B) endoribonucleases. C) polynucleotide phosphorylase. D) RNA polymerase II. E) small nuclear ribonucleoproteins (snRNPs).
E
The exchange of genetic material between homologous chromosomes
crossing over
Using Figure 10-3, which process would result in the formation of chromosome C from chromosome A and B?
crossing over
7. Which of the following statements about aminoacyl-tRNA synthetases is false? A) Some of the enzymes have an editing/proofreading capability. B) The enzyme attaches an amino acid to the 3' end of a tRNA. C) The enzyme splits ATP to AMP + PPi. D) The enzyme will use any tRNA species but is highly specific for a given amino acid. E) There is a different synthetase for every amino acid.
D) The enzyme will use any tRNA species but is highly specific for a given amino acid.
4. Which one of the following statements about ribosomes is true? A) The large subunit contains rRNA molecules; the small subunit does not. B) The RNA in ribosomes plays a structural, not catalytic, role. C) There are about 25 ribosomes in an E. coli cell. D) There are two major ribosomal subunits, each with multiple proteins. E) Ribosomes are relatively small, with molecular weights less than 10,000.
D) There are two major ribosomal subunits, each with multiple proteins.
Which of the following statements is false? In vitro, the strand-exchange reaction: A) can include formation of a Holliday intermediate. B) is accompanied by ATP hydrolysis. C) may involve transient formation of a three- or four-stranded DNA complex. D) needs RecA protein. E) requires DNA polymerase.
E
The entire collection of genes within human cells is referred to as the
human genome
When an area of a chromatid is exchanged with the matching area on a chromatid of its homologous chromosome, ___ occurs.
crossing over
1. Which of the following is a feature of site-specific recombination? A) A specific recombinase enzyme is required. 3 B) The energy of the phosphodiester bond is preserved in covalent enzyme-DNA linkage. C) Recombination sites have non-palindromic sequences. D) Insertions or deletions can result from site-specific recombination. E) All above.
E
2. The phosphodiester bonds that link adjacent nucleotides in both RNA and DNA: A) always link A with T and G with C. B) are susceptible to alkaline hydrolysis. C) are uncharged at neutral pH. D) form between the planar rings of adjacent bases. E) join the 3' hydroxyl of one nucleotide to the 5' phosphate of the next
E
20. In what location do you expect to find single-stranded siRNA? A) Attached to the major groove of DNA by hydrogen bonds B) Bound to an operon Repressor site C) Bound to RNA polymerase D) Bound to the small ribosomal subunit E) Bound to mRNA
E
3. Chargaff's rules state that in typical DNA: A) A = G. B) A = C. C) A = U. D) A + T = G + C. E) A + G = T + C.
E
6. Which of the following statements about the tRNA that normally accepts phenylalanine is false? (mRNA codons for phenylalanine are UUU and UUC.) A) It interacts specifically with the Phe synthetase. B) It will accept only the amino acid phenylalanine. C) It's about 80-nucleotides long. D) Phenylalanine can be specifically attached to an —OH group at the 3' end. E) The tRNA must contain the sequence UUU.
E) The tRNA must contain the sequence UUU.
17. By adding SDS (sodium dodecyl sulfate) during the electrophoresis of proteins, it is possible to: A) determine a protein's isoelectric point. B) determine an enzyme's specific activity. C) determine the amino acid composition of the protein. D) preserve a protein's native structure and biological activity. E) separate proteins exclusively on the basis of molecular weight.
E) separate proteins exclusively on the basis of molecular weight.
4. In the Watson-Crick model of DNA structure: A) both strands run in the same direction, 3' → 5'; they are parallel. B) phosphate groups project toward the middle of the helix, where they are protected from interaction with water. C) T can form three hydrogen bonds with either G or C in the opposite strand. D) the distance between the sugar backbone of the two strands is just large enough to accommodate either two purines or two pyrimidines. E) the distance between two adjacent bases in one strand is about 3.4 Å
E
5. In living cells, nucleotides and their derivatives can serve as: A) carriers of metabolic energy. B) enzyme cofactors. C) intracellular signals. D) precursors for nucleic acid synthesis. E) All of the above.
E
DNA upstream of the replication fork is unwound by a class of enzymes called what?
DNA helicases
Analysis of cloned genes - DNA sequencing
Dideoxynucleotide sequencing - has an -H not an -OH at the 3' end so it terminates replication - no where to form phosphodiester bond to the next nucleotide - YOU NEED .... -DNA polymerase -DNA template -primer -dATP, dGTP, dCTP, dTTP -small amount of a didNTP that is labeled 1. primer is annealed to DNA template 2. Then reaction mixture with DNA poly, all the bases, and the ddNTP (tagged with fluorescent dyes, each base has a diff color that this is attached to AKA ddATP - pink and ddGTP is green etc) 3. Eventually there will be a ddNTP inserted at every location so that each strand differs in length by 1 nucleotide because during primer extension it sometimes puts in ddNTP instead of dNTP 4. the products of this are added to a single lane on a capillary gel electrophoresis, and the bands are read by a detector system 5. the sequence is found by the extension of the primer, and is read from new strand not template strand
The _____________ recognize Ori C sequence to initiate DNA replication in E. coli
DnaA
DNA from an organism is cut by a restriction enzyme and so is the vector DNA, both cut by the same enzyme, and then they can base pair with their complementary tails, and then gaps are sealed with DNA ligase THIS MAKES RECOMBINANT DNA
Done in a test tube at low temperature, this is necessary for hydrogen bonds to stabilize and stick together. Ligase seals the phosphodiester bonds and now they're bonded from two different sources
Repair pathways: Ap endonuclease UvrAF, C, D protein mutS-mutL complex Translesion DNA polymerase DNA photolyase
Enzymes BER NER MMR translesion synthesis Direction repair
Santa Gertrudis cattle have the combined traits of good beef production and hot weather tolerance as a result of selection and INBREEDING.
False, hydridization
Many flowering plants such as roses, African violets, and orchids have been produced by the process of TEST CROSSING.
False, selective breeding
The hyp that genetic code consisted of groups of 3 nulceotides rather than groups of 2 nuclotides supports
Every amino acid has at least 1 corresponding codon
7. Many DNA polymerases have a proofreading 5' to 3' exonuclease.
F; 3 to 5
4. The ratio A+T/G+C is constant for all natural DNAs.
F; A+G=T+C
3. In the Watson-Crick model for the DNA double helix, there are two equally sized grooves that run up the sides of the helix
F; major and minor grooves are different in size
E. coli DNA polymerase III can initiate replication without a primer
F; needs a primer
5. In the Watson-Crick model of DNA structure: both strands run in the same direction, 5' to 3'; they are parallel.
F; opposite direction, anti parallel
Law and Regulation of GMOs
FDA/EPA/USDA - GM crops must be shown (by the manufacturer) to be "substantially equivalent" to nonGM versions of the crop. Toxicity and allergenicity of the crop are assessed based on the nature of the protein product of the genes that have been inserted. If they are deemed safe US law does not require labeling
in 1990, Young survivor of ADA deficiency: treated at age 4 with transformed T-cells -in 2000, 20 babies are Successfully treated For SCIDX but 3 of them get lymphoma
First done in 1990, and she got a correction, she got 4 or 5 treatments, and she got 25% of her immune system back, it is enough that she will survive -in 2000, 20 babies treated for SCID X they did 2 bone marrow replacements on these kids, and they had 95% success with this, most kids got 50% of their immune system back, then 3/20 of he kids got cancer, and 2 were treated and 1 died, and in these kids this virus put DNA where regulation of cells was important thus causing cancer -in 2002 human gene therapy stopped to get around this problem
Which of the following sequences is a palindrome and could be a restriction enzyme recognition site? >GAATTC> <CTTAAG<
GAATTC
___ is an application of the Human Genome Project that involves the insertion of normal genes into cells with defective genes in an attempt to correct genetic disorders.
Gene therapy
In 1974, Stanley Cohen and Herbert Boyer inserted a gene from an African clawed from into a bacterium. The bacterium produced the protein coded for by the inserted frog gene. The insertaion of a small fragment of frog DNA into the DNA of another species can most accurately be called
Genetic engineering
which of the following significantly contriubtes to the specificity of bp in double stranded DNA
H bonding b/w bases
Cloning and sequencing of genes is providing unexpected information on how genes function and evolve
Human Genome Project: Human - "finished" 2003 Drosophila -- done 1999 Caenorhabditis (worm) -- done 1997 Arabidopsis (plant) -- completed 2000 Yeast -- done 1996 E. coli -- done 1995 Mouse -- 2004 *Sequenced more than just humans - these different genomes are cross informative - more info about human genome by looking at other organisms than just humans alone -everything has been sequenced at this point - all in genbank
In Most Eukaryotes much of the Genome is repeated DNA sequences
Humans: 3 billion base pairs of DNA -50% repeated noncoding sequences: *most are transposable elements -45% single copy, noncoding sequences -5% codes for genes - this is the total of introns and exons (1.5% exons) ***L1 and Alu are the major transposable elements in the human genome
10. In bacteria the elongation stage of protein synthesis does not involve: A) aminoacyl-tRNAs. B) EF-Tu. C) GTP. D) IF-2. E) peptidyl transferase.
IF-2
Which of the structures in Figure 11-1 are composed of RNA?
II and IV
What is the genotype of generation 1 in Figure 10-5?
Ii
Which of the following statements correctly describes the role of telomerase in eukaryotic cells
It synthesizes new DNA a the 3' end of the chromosome
a ___________________ show the relative location of genes on a chromosome
Linkage Map
How should the top row of Figure 10-7 be read?
MMXX, MMXx, MmXX, MmXx
Genome sequencing based on mapped "contigs" (overlapping genomic DNA clones)
Map-based Cloning: 1. large DNA clones are mapped 2. individual contigs are restriction mapped subcloned and subclones are sequenced (subclones derived from one clones, and then that subclones can be cut up by restriction enzymes and mapped to determine its sequence) -first an entire chromosome is cut into short over lapping sequences most likely by restriction enzymes, -overlapping clones are mapped with in situ hybridization -subclones derived from one of many cloned fragments -each subclone can be characterized by restriction mapping, and this way each subclone will be cut at restriction sites, fragments cloned, and nucleotide sequence determined
Which of the following factors significantly contributes to the ability of eukaryotic cells to complete DNA replcation during the short window of time offered by S phase
Mulitiple origins of replciation are activated per chromosome
Adenine is to thymine as guanine is to
cytosine
which enzymes is directly responsible for sliding DNA along the surface of an assembled nuclosome to transferring a nuclosome form one DNa helix to another
Nucleosome- remodeling complexes
What genes cause cancer and why?
Oncogenic (cancer-causing ) Viruses carry oncogenes -DNA tumor viruses: Papilloma, Epstein Bar, Hepatitis B: Viral genes activate DNA replication in host -RNA tumor viruses - retroviruses: Infection by virus causes mis-expression of host genes -in humans RNA tumor viruses aren't a primary cause of cancer, because they don't directly cause cancer, but they may lead to mutations that do so cancer is a secondary affect of the virus -organisms other than human do have retroviruses that are the direct cause of cancer when the virus enters the cell and expresses its genes, it causes mis-expression of host genes
a technique that may be employed in the hUman Genome Projects is
PCR, automated gene sequencers, gel electrophoresis- All of these
10. Which one of the following statements about the elongation phase of protein synthesis is true? A) At least five high-energy phosphoryl groups are expended for each peptide bond formed. B) During elongation, incoming aminoacylated tRNAs are first bound in the P site. C) Elongation factor EF-Tu facilitates translocation. D) Peptidyl transferase catalyzes the attack of the carboxyl group of the incoming amino acid on an ester linkage in the nascent polypeptide. E) Peptidyl transferase is a ribozyme.
Peptidyl transferase is a ribozyme
A Ribosome contains three binding sites for tRNA molecules: the_______ or P-site, holds the tRNA molecule that is linked to the growing end of the polypeptide chain, and the _____-site, holds the incoming tRNA molecule that is charged with an amino acid and a E-site.
Peptidyl-site; Aminoacyl (A)
Vectors
Plasmids -origin of DNA replication -antibiotic resistance gene -unique restriction sites *Size of the recombinant fragment is limited to ~10 kb Vectors are bacterial plasmids- they're modified for cloning. The basic features of all of them is that they can replicate (origin of replication), antibiotic resistant genes (so you can introduce this without DNA cells recognizing it) and it must have places around the plasmid where restriction enzymes only cut once, and only once. You want it to be open up so it can accept an insert, and the plasmids have been modified where the sites only cut once. -the plasmid vector is removed from host cell and cut by a restriction enzyme, DNA to be cloned is cut with the same restriction enzyme, and then these 2 pieces are ligated, and introduced into bacterial host cells by transformation
A useful device for predicting the possible offspring of crosses between different genotypes is the ___.
Punnett square
During the process of transcription, DNA serves as the template for making ___, which leaves the nucleus and travels to the ribosomes.
RNA
Primases synthesize a short stretch of ______ to prime further synthesis
RNA
Translation is to protein as transcription is to ___.
RNA
The uniting of the male and female gametes
fertilization
Which one of the following components is not required for DNA synthesis
RNTPs ribo Triphos
In pea plants, inflated pods (R) are dominant to constricted pods (r). Which of the following crosses is a test cross between inflated pods and constricted pods?
RR x rr
In pea plants, inflated pods are dominant to constricted pods. Which of the following crosses is a test cross between inflated pods and constricted pods
RR x rr
___________are used to cleave DNA into fragments
Restriction Enzymes
___ are used to cleave DNA into fragments.
Restriction enzymes
B-form DNA in vivo is a ________-handed helix, _____ Å in diameter, with a rise of ____ Å per base pair
Right; 20; 3.4
Criticisms of GMOs
Safety -- Food Allergens Consumer choice - Labeling Bio Pollution Patents and control of food supply US decided that these crops must be substantially equivalent - have to show toxicity of protein product of the gene you put in (protein that dna codes for) -test to see if these proteins bring with them issues of toxicity or Allergenicity - that you can directly test -Bio pollution - biological polltuion - can these plants hybridize with the genes of native plants and then this can spread **For Example - with corn and cotton, there are no wild plants for the hybrid to cross with so there are no genes for it to hybridize with > worried that it can get away into wild plants -issue of patents and food supply - big companies that are producing seeds farmers are using > they have control over food supply
During DNA replication the parental DNA strands serve as templates for the synthesis of new progeny DNA stands, this is called
Semi-conservation
Which one of the following is true about the genetic code? A) All codons recognized by a given tRNA encode different amino acids. B) It is absolutely identical in all living things. C) Several different codons may encode the same amino acid. D) The base in the middle position of the tRNA anticodon sometimes permits "wobble" base pairing with two or three different codons. E) The first position of the tRNA anticodon is always adenosine.
Several different codons may encode the same amino acid.
PCR Uses DNA Polymerase to copy DNA in the test tube repeated rounds of replication
Specificity of PCR depends on choosing the right primers and the annealing Temp used
2. The DNA of virtually every cell is underwound (i.e., negatively supercoiled) relative to Bform DNA
T
which of the following is ture readign the dirveion of new DNA synthesis during relication
THe leading strand DNA polymerase synthesizes DNA in the same direction that the replication fork is moving
The most abundant sequences in the human genome are
TRANSPOSABLE ELEMENTS - more transposable elements and repeats occur in more complex organisms
Which of the following are features of the wobble hypothesis? A) A naturally occurring tRNA exists in yeast that can read both arginine and lysine codons. B) A tRNA can recognize only one codon. C) Some tRNAs can recognize codons that specify two different amino acids if both are nonpolar. D) The "wobble" occurs only in the first base of the anticodon. E) The "wobble" occurs only in the third base of the anticodon.
The "wobble" occurs only in the first base of the anticodon.
Using STRs to follow human history
The Y chromosome does not undergo recombination, so related males inherit identical STR alleles - haplotypes -Y chromosome is directly inherited from father to son - less genetic variability in Y chromosome than autosomal chromosomes (not sex) -even 2 unrelated males may have the same Y profile if they share a distant ancestor
White colonies carry a recombinant DNA plasmid Blue ones do not
The cells making beta G turn blue, and if they don't, they're white. The white colonies contain a plasmid with an insertion ability. The blue ones just have the plasmids by themselves. -Original plasmid has lac-z gene to make x-gal and form blue colonies > multiple cloning site is cut > DNA cut with the same restriction enzyme is added and ligased to plasmid > DNA frag inserted anywhere in multiple cloning site, the lac Z gene is disrupted and cannot make beta G > recombinant plasmid cannot metabolize x-gal and will form white colonies
Output of a DNA sequencing reaction using fluorescently labeled bases
The fluorescent tags on the detector and signal system: as you get to longer pieces the peaks go down and the widths become wider and then you can't distinguish the peaks anymore ***ALL OF THIS TECHNOLOGY CAME OUT OF HUMAN GENOME PROJECT
Whats interesting? Why do bacteria have them all?
These are sequences that cut DNA so youd expect it to cut its own DNA which would be lethal, but it makes restriction enzymes and a corresponding methlyase at the same time, which methylates the other strand which blocks cutting. This doesn't happen in humans, so these enzymes are for the bacteria's immune system. Foreign DNA is moved around and if the cell has the restiction enzyme it can cut the foreign DNA up and protect itself from being genetically modified. In the labs- we get rid of this function so DNA can be cloned
Why would relaxed samples migrate faster in gel
Treatment w/ ethidium relaxed the negatively supercoiled cccDNA and positively supercoiled the relaxed cccDNA
Southern Blot of DNA from 4 Species: Human, mouse, hamster, and cow -Cut with EcoRI or HindIII or PstI -Probed with a fragment of the human cystic fibrosis gene
This is called a zoo block - they use animal DNA and probed it with the human cystic fibrosis gene to see if it would hybridize - aka is their DNA complementary to our DNA sequence for cystic fibrosis gene? -The cow showed some hybridization to the human gene, yet not in the same places as the human -The mouse and hamster don't have any hybridization - showing a bigger evolution between us and hamsters/mouse than us and cows -When you have a piece of DNA from 1 species and hybridize it with another species the sequences aren't the same, so hybridization isn't the same
Human insulin (humulin) was the first human protein to be produced in bacteria using recombinant DNA
To make recombinant human insulin... -mature insulin in made of 2 polypeptide chains: A (21 amino acids) and B (60 amino acids) -synthetic genes to make A and B were made by oligonucleotide synthesis (63 nucleotides for A and 90 for B) -These oligonucleotide sequences are inserted into separate vectors at the tail end of a cloned E. coli lacZ gene -The recombinant plasmids were transformed into E. coli host cells, where the B-gal/insulin fusion protein was synthesized and accumulated in cells -B-gal/insulin fusion proteins were extracted from host cells and purified -insulin chains were released from B-gal by treating with cyanogen bromide -insulin subunits were purified and mixed to produce a functional insulin molecule
_______are produced when DNA from another species is inserted into the genome of an organism which then begins to produce the protein encoded on the recombinant DNA
Transgenic Organism
In 1974, Stanley Cohen and Herbert Boyer inserted a gene from an African clawed from into a bacterium. The bacterium produced the protein coded for by the inserted frog gene. The bacterium containing functional frog DNA would be classified as a ______
Transgenic organism
___ are produced when DNA from another species is inserted into the genome of an organism, which then begins to produce the protein encoded on the recombinant DNA.
Transgenic organisms
1.The linking number (Lk) of a closed-circular DNA molecule can be changed only by breaking one or both strands.
True
You have benefited from SELECTIVE BREEDING by having more agricultural produce than would have been possible otherwise.
True
Prenatal Testing: Amniocentesis ASO or FISH can be used for genetic diseases
Use amniotic fluid and obtain the fluid and the fetal cells from it - culture this on a pea tree dish and analyze it using recombinant DNA methods for looking at genetic and biochemical analysis -you can do it before insemination if you couple it with in vitro fertilization
A gene gun and a virus may both be classified as______ because they are mechanisms by which foreign DNA may be transferred into a host cell
Vectors
11. It is possible to convert the Cys that is a part of Cys-tRNACys to Ala by a catalytic reduction. If the resulting Ala-tRNACys were added to a mixture of (1) ribosomes, (2) all the other tRNAs and amino acids, (3) all of the cofactors and enzymes needed to make protein in vitro, and (4) mRNA for hemoglobin, where in the newly synthesized hemoglobin would the Ala from Ala-tRNACys be incorporated? A) Nowhere; this is the equivalent of a nonsense mutation. B) Wherever Ala normally occurs C) Wherever Cys normally occurs D) Wherever either Ala or Cys normally occurs E) Wherever the dipeptide Ala-Cys normally occurs
Wherever Cys normally occurs
What is the genotype in the bottom left-hand quadrant in Figure 10-6?
Ww
Tissue culture allow us to grow entire plants from fragments of stems, roots or leaves
You can genetically modify plants in tissue culture > make genetic modification of cells and take these cells to make a plant -make pieces of plant by manipulating culture conditions > you make roots and put them in soil and they'll grow **Easiest to make genetic modification is in tissue culture
Structure III in Figure 11-1 represents a(n) ___.
amino acid
Gene Families Evolve by Gene Duplication followed by Sequence Drift
a-globin versus b-globin 50% sequence similarity d-globin versus b-globin 93% same (d is on beta chain) -The globin genes (myoglobin + hemoglobin (a/b)) - the relationships between them and the points at which the gene duplication occurs - based on sequence similarities -You get the timeline by figuring out when the gene gets an amino acid substitution (rate of mutation assuming its constant) and another way is to use the fossil record, knowing where different groups diverged from each other to see what alpha genes they do and don't have -We think these dates are accurate, divergences in gene families don't occur often but over time
Wind, humidity, and rocks are all ___ in a terrestrial ecosystem.
abiotic factors
A cell moves particles from a region of lesser concentration to a region of greater concentration by ___.
active transport
The Human Genome Project may make use of which of the following to diagnose genetic disorders before birth?
all of the above
The causes of cancer may include which of the following?
all of the above
What would be the result of the test cross in Figure 13-4 if the unknown were homozygous long ears?
all of the offspring would have long ears
What would be the result of the test cross in fig 13-4 of the unknown were homozygous long ears
all of the offspring would have long ears
A technique that may be employed in the Human Genome Project is ___.
all of these
Which of the following are applications of genetic engineering?
all of these
Genes located on homologous chromosomes may have alternate forms that control different forms of a trait.
alleles
The chromosomes shown in Figure 8-6 are in which state of mitosis?
anaphase
The phase of mitosis in which the sister chromatids separate from each other is
anaphase
The Neanderthal Genome has been sequenced
ancestors evolve into neanderthals 350,000-500,000 years ago - and they died out 30,000 years ago and modern homo sapiens take over -All non-African modern humans have 2-4% Neanderthal DNA sequences
The _______ in a tRNA molecule is designed to base-pair with a complementary sequence of three nucleotides, the ___________ in an mRNA molecule.
anti-codon; codon
In which part of the cell does this process shown in Figure 11-1 take place?
at the ribosomes
manufactures food using energy from the sun or from chemical compounds
autotroph
group formed by several populations
biological community
To explain and show how the amount of living materials at each trophic level of a food chain changes, you could use a pyramid of ___.
biomass
Ecosystems, biotic factors, and abiotic factors make up the ___.
biosphere
layer of Earth that supports life
biosphere
In a pond ecosystem, ducks, mosquitoes, pond plants, and frogs are ___ factors.
biotic factors
Which of the following explains why a cell's size is limited?
both a and c
cDNA = "complementary DNA" made from mRNA
cDNA synthesis - uses Reverse Transcriptase to make a DNA copy of mRNA 1) mix mRNAs with oligo-dT primers (single stranded) and these bind to the poly A tail. oligo primer is (TTTTT) 2) reverse transcriptase extends the oligo-dT primer and makes a complementary DNA copy of the mRNA 4) The RNA part of the hybrid (because now you have one strand of DNA and 1 RNA) is partially digest by RNAse H, the 3 ends of remaining RNA serves as primers for DNA poly 1, to make the second strand of DNA 5) and the opposing strand of DNA is made with DNA poly 1 and ligase to make the final double stranded cDNA molecule
The uncontrolled division of cells that results in a malignant growth is known as
cancer
15. Prosthetic groups in the class of proteins known as glycoproteins are composed of: A) carbohydrates. B) flavin nucleotides. C) lipids. D) metals . E) phosphates.
carbohydrates
for the diagnosis of a genetic disorder, many cells are required, but only a few need to be taken from the individual. These cells are grown in _____________so that enough DNA can be obtained to run the necessary tests
cell culture
The Human Genome Project may make use of which of the following to diagnose genetic disorders before birth?
cell cultures, PCR, automated gene sequencers---All of the above
The sequence of growth and division of a cell makes up the
cell cycle
Which series is arranged in order from largest to smallest in size?
cell, nucleus, chromosome, DNA, nucleotide
Unlike plant cells, animal cells contain ___.
centrioles
A chromatid is attached to a spindle fiber by the ___.
centromere
The structures that hold together sister chromatids are
centromeres
The dark-staining structures that carry the genetic material are the
chromosomes
Recombinant DNA are currently used to produce
clothing dye, cheese, and laundry products
Recombinant DNA are currently used to produce ___.
clothing dye, cheese, and laundry products
Each set of three nitrogen bases representing an amino acid is referred to as a(n)
codon
relationship between organisms in which one organism benefits and the other is neither harmed nor benefited
commensalism
An ecologist who studies how several species in an area interact is interested in the biologist organization called a(n) ___.
community
Nitrogen is released to the abiotic parts of the biosphere from the processes of death and ___.
decay by bacteria
breaks down dead organisms
decomposer
Before plants can reuse many organic materials, the materials must be broken down by ___.
decomposers
Water, carbon, and nitrogen are released back into the atmosphere during ___.
decomposition
A DNA nucleotide may be made up of a phosphate group, along with ___.
deoxyribose sugar and thymine
Which of the following would be an example of gene therapy technology?
development of a nasal spray that contains copies of the normal gene that is defective in persons with cystic fibrosis
which of the following would be an example of gene therapy technology
development of a nasal spray that contains copies of the normal gene that is defective in persons with cystic fibrosis
movement of particles from an area of higher concentration to one of lower concentration
diffusion
A cross involving two different traits
dihybrid
Cells containing two alleles for each trait are described as ___.
diploid
Watson and Crick called the three-dimensional shape of DNA a
double helix
Watson and Crick, with the help of Rosalind Franklin, developed the ___ model of DNA.
double helix
study of how living things relate to each other and to their environment
ecology
In ecological classification, the next smallest level after the biosphere is the ___.
ecosystem
Organism, population, and community make up the ___.
ecosytem
If the sides of a cell double in length, its volume increased by ___ times.
eight
Which of the following is NOT a form of passive transport?
endocytosis
Glo Fish and Glo Pigs!
engineered to express the gene for green fluorescent protein cloned from jelly fish
Restriction maps
establishes the number of, order of, and distances between restriction sites along a cloned segment of DNA > thus providing information about the length of the cloned gene and the location of restriction sites within the clone.
release of wastes of cell products from inside to outside a cell
exocytosis
What cell process is responsible for the effect shown in Figure 8-5?
facilitated diffusion
simple model for showing how matter and energy move through an ecosystem
food chain
Tropic level and food chain are parts of a ___.
food web
network of interconnected food chains
food web
If the sides of a cell double in length, its surface area becomes ___ times as large.
four
A(n) ___ involves the addition or deletion of a single base in a DNA molecule.
frameshift mutation
The ___ produced by each parent are shown along the sides of a Punnett square.
gametes
The process used to separate DNA segments of different lengths is
gel electrophoresis
The process used to separate DNA segments of different lengths is ___.
gel electrophoresis
a segment of DNA that controls the production of a protein
gene
The Human Genome Project has involved sequencing and mapping the human genome. The most important benefit of this information has been the diagnosis of genetic disorders. Once a genetic disorder is diagnosed, ___ can be used as a possible treatment.
gene therapy
The human Genome Project has involved sequencing and mapping the human genome. The most important benefit of this information as been the diagnosis of genetic disorders. Once a genetic disorder is diagnosed, ______ can be used as a possible treatment
gene therapy
_______is an application of the Human Genome Project that involves the insertion of normal genes into cells with defective genes in an attempt to correct genetic disorders
gene therapy
In 1974, Stanley Cohen and Herbert Boyer inserted a gene from an African clawed frog into a bacterium. The bacterium produced the protein coded for by the inserted frog gene. This insertion of a small fragment of frog DNA into the DNA of another species can most accurately be called ___.
genetic engineering
Crossing over results in a ___.
genetic recombination
The intergenic parts of the human genome consist of which types of DNA sequences
genome-wide repeats, microsatellite DNA and regulatory sequences
Which one of the following nucleotide pair bonds would be found in a DNA molecule?
guanine-cytosine
Sea stars live in saltwater ecosystems. Some species live in shallow tidal pools, while others live in the deepest parts of the oceans. This is a description of the ___ of sea stars.
habitat
place where an organism spends its life
habitat
A cell that contains one member of each chromosome pair
haploid
Energy that passes through a food chain is lost to the environment as ___.
heat
The passing of characteristics from parents to offspring
heredity
The passing on of traits from parents to offspring is called ___.
heredity
Omnivores, carnivores, herbivores, scavengers, and decomposers are all
heterotroph
obtains energy and nutrients from autotrophs
heterotroph
What is the genotype of the unknown rabbit in Figure 13-4?
heterozygous
What is the genotype of the unknown rabbits
heterozygous
Which of the following describes an organism that has the genotype Bb?
heterozygous
Assembly of nucleosomes requires
histone "chaperones"
in add'l to histon composition and chromatin-remodeling, what is the 3rd way that the chromation structure can be regulated?
histone modification
many crop plants such as wheat and corn have been developed as _______in order to develop larger and stronger plants
hybrids
Water moves into a cell placed in a(n) ___ solution.
hypertonic
Water moves out of a cell if the cell is placed in a(n) ___ solution.
hypertonic
the concentration of dissolved substances outside the cell is higher than the concentration inside the cell
hypertonic solution
__________ is used to develop pure breeds
inbreeding
As matter and energy move from grasses to coyotes, the amount of available energy ___.
increases or decreases but population size remains the same
Which of the following rxns w/ covalently closed circular DNA does NOT result in a new lk
incubation w/ ethidium
The statement: "In meiosis, the way in which a chromosome pair separates does not affect the way other pairs separate," is another way of expressing Mendel's law of ___.
independent assortment
What is the phenotype of generation 1 in Figure 10-5?
inflated
The processivity of a DNA polymerase depends on all of the following
interaction b/w the thumb domain of DNA polymerase and the DNA 2. associateion of the DNA polymerase w/ a sliding clamp 3. interaction b/w minor goorve of DNA and the palm domain of the DNA polymerase
12. The chirality of an amino acid results from the fact that its α carbon: A) has no net charge. B) is a carboxylic acid. C) is bonded to four different chemical groups. D) is in the L absolute configuration in naturally occurring proteins. E) is symmetric.
is bonded to four different chemical groups
the concentration of dissolved substances in the solution is the same as the concentration of dissolved substances inside the cell
isotonic solution
DNA binding proteins primarily recongize a specific seqof double stranded DNA through
itneractions w/ the exposed chemical groups in the major groove of the DNA
Recombinant DNA
joining of DNA from different organisms in the test tube •isolation of individual genes (Gene Cloning) •study of gene function and regulation of expression •cloned genes can be sequenced and their base sequences can be modified •transgenic organisms - insertion of cloned genes into animals, plants or bacteria "GENETIC ENGINEERING"
An Okazaki fragment is a segment of DNA that is an intermediate in the synthesis of the leading strand
lagging strand
Evolutionary rln b/w complexity of organisms and gene density
less complex organisms have increased gene density
By the end of prophase, each of the following has occurred except ___.
lining up of chromosomes in the cell
A(n) ___ shows the relative location of genes on a chromosome.
linkage map
A dog's phenotype can be determined by ___.
looking at the dog
An application of using DNA technology to help environmental scientists would be
make transgenic bacteria that can be used to clean up oil spills more quickly than do the natural bacteria
An application of using DNA technology to help environmental scientists would be ___.
make transgenic bacteria that can be used to clean up oil spills more quickly than do the natural bacteria
The type of cell division that produces gametes
meiosis
Energy flows from ___.
mice to cats
Which of the following monitors a cell's progress from phase to phase during the cell cycle?
microtubules
Among the following, the term that includes the other is ___.
mitosis
The process by which nuclear material is divided equally between two new cells is
mitosis
The numbers in Figure 10-1 represent the chromosome number found in each of the dog cells shown. The processes that are occurring at A and B are ___.
mitosis and fertilization
When parts of chromosomes are broken off and lost during mitosis or meiosis, the result is a(n)
mutation
X rays, ultraviolet light, and radioactive substances that can change the chemical nature of DNA are classified as ___.
mutegens
Both the alga and the fungus are benefited from their relationship in a lichen. This relationship is one of ___.
mutualism
Some birds are known as honey guides because they may be followed by humans to wild beehives. When the humans take honey from the hives, the birds are able to feast on the honey and bees, too. This type of relationship can BEST be described as ___.
mutualism
What will be the result of the mutation in Figure 11-4?
nearly every amino acid in the protein will be changed
The hershey and case exp using S and P labeled T2 bacteriophage supported the hyp that DNA is the genetic material b/c
new T2 progeny contained P-labeled DNA
Cougars are predators that often eat weakened or diseased animals. This is a description of the ___ of cougars.
niche
Mendel's law of segregation states that during meiosis, the factors that control each trait separate, and only ___ from each fair is/are passed to the offspring.
one factor
What level of organization is shown in Figure 8-7?
organ system
Which of the following structures is the most complex?
organ system
If a cell is placed in salt water, water leaves the cell by ___.
osmosis
diffusion of water molecules through a selectively permeable membrane
osmosis
relationship between organisms in which one organism benefits at the expense of another
parasitism
If cells are placed in a strong sugar solution, water will ___.
pass from the cells to the sugar solution.
It enciricles one of the 2 DNA strands upstream of the fork and uses the energy provided by ATP hydrolysis to separate the strands, The unwinding of the DNA introduces ______ into the double strandedDNA upstream to the fork that need to be removed by topoisomerase
positive supercoils
The backbone of a DNA molecule is made of which two components?
phosphate molecules and deoxyribose sugars
bonds that link adjacent nucleotides in both RNA and DNA
photodiester
Carbon dioxide in the atmosphere enters the biotic parts of the biosphere through ___.
photosynthesis
The structure most responsible for maintaining cell homeostasis is the ___.
plasma membrane
A(n) ___ is a small ring of DNA found in a bacterial cell.
plasmid
Multiple cloning site
plasmid vectors have this which allows them to have many restriction sites for many restriction enzymes- allows scientist to clone a range of different fragments generated from many common restriction enzymes
A DNA segment is changed from-AATTAG- to -AAATAG-. This is a ___.
point mutation
A small amount of DNA obtained from a mummy or from frozen remains of a human may be cloned. In order to clone small amounts of DNA, ___ needs to be used to generate larger quantities of the DNA.
polymerase chain reaction techniques
A small amount of DNA obtained from a mummy or from frozen remains of a human may be cloned. In order to clone small amounts of DNA, ____needs to be used to generate larger quantities of the DNA
polymerase chain reaction techniques
How to make cDNA? Use reverse transcriptase- turns RNA to DNA, and the source of this is retroviruses. The retroviruses are RNA viruses that replicate by turning their genome into DNA and inserting it into genomes, (HIV) To clone a mRNA for eurkaryote—isolate total RNA from the cell, add to it a short piece 20 bases long of just T's (oligo dt primer) and they will base pair with the poly A tail. That serves as a primer site for reverse transcriptase site, and it will start the replication . RNA and DNA together now, and RNA H degrades RNA and it only degrates RNA that is attached to DNA. You add this in to take away the RNA but not all, then add DNA poly 1 and it will fill in the spaces and will use the primer to chug along and cut out RNA and replace it with DNA. Add DNA ligase & seal, now you have a DNA piece that coresponds with a messanger. To get into a plasmid, you must manipulate more steps (don't need to know) Just know reverse transcriptase & enzyme Rnase H For HIV- it has RNAse into it (read)
powerpoint notes
The relationships between the cats and mice could best be described as ___.
predator-prey
in vitro DNA synthesis requires DNA polymerase DNTPsand a what junction
primer/templcate
The longest phase of the cell cycle is ___.
prophase
A gene is a segment of DNA that controls the production of ___.
proteins
The process by which a DNA molecule is copied is called ___.
replication
16. For the study of a protein in detail, an effort is usually made to first: A) conjugate the protein to a known molecule. B) determine its amino acid composition. C) determine its amino acid sequence. D) determine its molecular weight. E) purify the protein.
purify the protein
Peptide bond formation is catalyzed by a catalytic activity that is thought to be mediated by the ________ molecule in the large ribosomal subunit
rRNA
Ribosomes are made of ___.
rRNA and protein
According to Figure 10-5, the constricted pod shape is ___.
recessive
The process by which DNA makes a copy of itself is called
replication
feeds on dead organisms
scavenger
Many flowering plants such as roses, Aftrican violets, and orchids have been produced by the process of __________________
selective breeding
You have benefited from ______________ by having more agricultural produce than would have been possible otherwise
selective breeding
Gel electrophoresis is a technique used to
separate DNA fragments by charge and length
Gel electrophoresis is a technique used to ___.
separate DNA fragments by charge and length
The law of independent assortment states that the inheritance of alleles for a different trait if the genes for the traits are on ___.
separate chromosomes
DNA is to RNA as double stranded is to
single stranded
In a dividing cell, the football-shaped structure consisting of thin fibers is the
spindle
The historical method used to assign genes to particular human chromosome was to
study linkage data from human pedigrees
The historical method used to assign genes to particular human chromosome was to ___.
study linkage data from human pedigrees
Applicants for the second job of the Help Wanted ad in Table 11-1, "Accuracy and Speed," could qualify if they were ___.
tRNA
The molecule ___ brings amino acids to the ribosomes for the assembly of proteins.
tRNA
Northern blotts are also used to study splicing reactions - like in this case ^, the splicing is done differently in the heart and the brain because they have different amount of RNA
tells us about alternative splicing, size of mrna transcript, and estimate relative transcriptional activity of a gene
To determine if an individual with a dominant phenotype is homozygous or heterozygous, a ___ is used.
test cross
To determine if an individual with a dominant phenotype is homozygous or heterozygous, a ____________ is used
test cross
19. The term specific activity differs from the term activity in that specific activity: A) is measured only under optimal conditions. B) is the activity (enzyme units) in a milligram of protein. C) is the activity (enzyme units) of a specific protein. D) refers only to a purified protein. E) refers to proteins other than enzymes.
the activity in a mg of protein
Eukaryotic Genomes Vary Greatly in Size
the bigger the genome, the more repeated sequences there are
Genome
the complete set of DNA in a single cell of an organism
Which of the following would be least likely to happen as a result of a mutation in a person's skin cells?
the person's offspring have mutated skin
Why would the absence of DNA primers in PCR cause the rxn to fail
the rxn lacks a primer: template junction and thus a free 3/'OH group when primers are absent
Examine the pieces of DNA represented in Fig 13-1. Why are the nucleotide sequences on both strands referred to a palindromes
the sequences are the same but run in opposite directions
Examine the pieces of DNA represented in Figure 13-1. Why are the nucleotide sequences on both strands referred to as palindromes?
the sequences show chromosome mutation
Watson and Crick were the first to suggest that DNA is ___.
the shape of a double helix
13. At the isoelectric pH of a tetrapeptide: A) only the amino and carboxyl termini contribute charge. B) the amino and carboxyl termini are not charged. C) the total net charge is zero. D) there are four ionic charges. E) two internal amino acids of the tetrapeptide cannot have ionizable R groups.
the total net charge is zero
Pollination can best be described as ___.
the transfer of the male pollen grain to the female organ
Each bacterial colony is a clone, carrying a specific cloned DNA sequence
the whole thing plated on the medium is a colony, and each are a clone of some specific DNA sequence
If cancer is present, what is the likely explanation for what happened to cells B and D?
they died off because the cancerous cells deprived them of nutrients
Cells that work together to perform the same function are organized into
tissues
Messenger RNA is formed in the process of ___.
transcription
Which of the following are applications of genetic engineering?
transgenic bacteria in agriculture, transgenic plants and animals, transgenic bacteria in industry- all of these
In 1974, Stanley Cohen and Herbert Boyer inserted a gene from an African clawed frog into a bacterium. The bacterium produced the protein coded for by the inserted frog gene. The bacterium containing functional frog DNA would be classified as a ___.
transgenic organism
The process illustrated in Figure 11-1 is called ___.
translation
The chromosome abnormality that occurs when part of one chromosome breaks off and is added to a different chromosome is ___.
translocation
Water is lost to the abiotic parts of the biosphere from the biotic parts by the process of ___.
transpiration
step in the passage of energy and matter through an ecosystem
trophic level
A gene gun and a virus may both be classified as ___ because they are mechanisms by which foreign DNA may be transferred into a host cell.
vectors
As a cell grows, its ___ increases more than its ___.
volume, surface area
The ___ consists of evaporation, precipitation, transpiration, runoff, and respiration.
water cycle
proto-oncogenes encode transcription factors that stimulate expression of other genes, signal transduction molecules that stimulate cell division, and cell cycle regulators that move cells through the cycle
when a proto-oncogene is mutated or overly expressed and contributes to the development of cancer it is an ***ONCOGENE***
Mutations in the EGF (epidermal growth factor) and neu growth factor RECEPTORS result in stimulation of cell division even in the absence of the hormone
when it loses the functional growth factor receptors, it causes it to not need the hormone to bind to cause cell division, so it is always active -some mutations cause it to bind the hormone and dimerize it doubling the output of the cell division -mutation of the neu growth factor receptor is a base substitution -mutation in EGF receptor is a deletion
The cell produced when a male gametes fuses with a female gamete
zygote
