Suppose that you wanted to study retroviral integration mechanisms by determining the nucleotide sequence at the integration site-sev- eral dozen nucleotides on each side of the viral-cellular DNA junc- tion. Describe how to isolate DNA containing a junction site in amounts sufficient for sequence analysis.
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A: A protospacer adjacent motif (PAM) is a 2–6 base pair DNA sequence tareted by Cas9 nuclease.
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A: Introduction Deoxyribonucleic acid (DNA) is a polymer made up of two polynucleotide chains that coil…
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A: Processivity is the average number of bases a polymerase enzyme will extend and incorporate base…
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A:
Q: You are cloning the genome of a new DNA virus into pUC18. You plate out your transformants on…
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Deoxyribonucleic acid (DNA) is a molecule composed of two polynucleotide chains that coil around each other to form a double helix carrying genetic information for the development, functioning, growth of all organisms.
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- Assume that a viral DNA molecule is a 50@μm-long circularstrand of a uniform 2-nm diameter. If this molecule is containedin a viral head that is a 0.08@μm-diameter sphere, will the DNAmolecule fit into the viral head, assuming complete flexibility ofthe molecule? Justify your answer mathematically.The anti-viral drug Acyclovir is a nucleotide analog that is lacking the 3’ OH group which is required to form a 3’→5’ phosphodiester bond. This drug is ineffective against DNA polymerases with proofreading abilities, which is why human DNA polymerases are not targeted. Acyclovir can be used to treatsevere cases of Epstein-Barr viral (EBV) infection, but has little to no effect under non-severe infections. Based on this information, EBV will use ________ DNA polymerase during severe infections and __________ DNA polymerase during non-severe infections. Human; Human EBV; Human EBV; EBV Human; EBVIn a bacterial culture in which all cells are unable to synthesizeleucine (leu-), a potent mutagen is added, and the cells areallowed to undergo one round of replication. At that point, samplesare taken, a series of dilutions are made, and the cells areplated on either minimal medium or minimal medium containingleucine. The first culture condition (minimal medium) allowsthe growth of only leu+ cells, while the second culture condition(minimal medium with leucine added) allows growth of all cells.The results of the experiment are as follows: Culture Condition Dilution ColoniesMinimal medium 10-1 18Minimal medium + leucine 10-7 6What is the rate of mutation at the locus associated with leucinebiosynthesis?
- A biological motor used to package double-stranded (ds) DNA into viral capsids is able toexert 50 pN of force. A. If the active site has a surface area of 10^-17 m2 , to what pressure can the biological motor package dsDNA within the viral capsid before stalling? B.If the viral capsid has a radius of 30 nm, how much potential energy is stored due to the pressure-confinement of the dsDNA within the capsid? C. Compare this to thermal energy at room temperature, kBT, where kB=1.38*10^-23 J/K is the Boltzmann constant.In a typical microbiology laboratory, reasons for no bands from a gel of a polymerase chain reaction may bedue to errors relating to omission of ingredients in the reaction mix and absence of the target sequence inthe template DNA. Based on (i) primer problem and (ii) purity/potential contamination of the target sequence, explain the reason for non-appereance on bands."DNA polymerase I serves a secondary function in vivo,now believed to be critical to the maintenance of fidelityof DNA synthesis" Explain this statement ?
- . The double-stranded circular DNA molecule thatforms the genome of the SV40 tumor virus can be denatured into single-stranded DNA molecules. Becausethe base composition of the two strands differs, thestrands can be separated on the basis of their densityinto two strands designated W(atson) and C(rick). When each of the purified preparations of the single strands was mixed with mRNA from cells infectedwith the virus, hybrids were formed between the RNAand DNA. Closer analysis of these hybridizationsshowed that RNAs that hybridized with the W preparation were different from RNAs that hybridized withthe C preparation. What does this tell you about thetranscription templates for the different classes ofRNAs?A number of yeast-derived elements were added to thecircular bacterial plasmid pBR322. Yeast that requireuracil for growth (Ura− cells) were transformed withthese modified plasmids and Ura+ colonies were selected by growth in media lacking uracil. For plasmidscontaining each of the elements listed in parts (a) to(c), indicate whether you expect the plasmid to integrate into a chromosome by recombination, or insteadwhether it is maintained separately as a plasmid. If theplasmid is maintained autonomously, is it stably inherited by all of the daughter cells of subsequent generations when you no longer select for Ura+ cells (that is,when you grow the yeast in media containing uracil)?a. URA+ geneb. URA+ gene, ARS c. URA+ gene, ARS, CEN (centromere)d. What would need to be added in order for these sequences to be maintained stably in yeast cells as alinear artificial chromosome?The modular nature of eukaryotic activator proteinsgave scientists an idea for a way to find proteins thatinteract with any particular protein of interest. Theidea is to use the protein–protein interaction to bringtogether a DNA-binding region with an activation region, creating an artificial activator that consists oftwo polypeptides held together noncovalently by theinteraction.The method is called the yeast two-hybrid system,and it has three components. First, the yeast contains areporter gene construct in which UASG (an enhancerlike sequence that binds the activator Gal4 as describedin Problem 8) drives the expression of an E. coli lacZreporter (encoding the enzyme ß-galactosidase) from ayeast promoter. Second, the yeast also expresses a fusion protein in which the DNA-binding domain of Gal4is fused to the protein of interest; this fusion protein iscalled the bait. The third component is a cDNA librarymade in plasmids, where each cDNA is fused in frameto the activation domain of…
- Dyskeratosis congenita (DKC) is a rare human genetic disorderaffecting telomere replication. Mutations in thegenes encoding the protein or RNA subunits of telomeraseresult in very short telomeres. DKC symptoms include bonemarrow failure (reduced production of blood cells) and anemia.If symptoms are severe, a bone marrow transplant may be theonly form of effective treatment. In one case, clinicians recommendedthat a 27-year-old woman with a dominant form of DKCundergo a bone marrow transplant to treat the disorder. Her foursiblings were tested, and her 13-year-old brother was identifiedas the best immunologically matched donor. However, beforebeing tested, he was emphatic that he did not want to knowif he had DKC. During testing, it was discovered that he hadunusually short telomeres and would most likely develop symptomsof DKC. Although the brother is an immunologically matched donor for his sister, it would be unethical for the clinicians to transplant bone marrow from the brother to…On further analysis of the DNA described in conceptual questionC21, you discover that the triplex DNA in this alien organism iscomposed of a double helix with a third strand wound within themajor groove (just like the DNA in Figure shown). How would youpropose that this DNA is able to replicate itself? In your answer,be specific about the base-pairing rules within the double helixand which part of the triplex DNA would be replicated first.Dyskeratosis congenita (DKC) is a rare human genetic disorderaffecting telomere replication. Mutations in the genesencoding the protein or RNA subunits of telomerase resultin very short telomeres. DKC symptoms include bone marrow failure(reduced production of blood cells) and anemia. If symptoms aresevere, a bone marrow transplant may be the only form of effectivetreatment. In one case, clinicians recommended that a 27-yearoldwoman with a dominant form of DKC undergo a bone marrowtransplant to treat the disorder. Her four siblings were tested, andher 13-year-old brother was identified as the best immunologicallymatched donor. However, before being tested, he was emphaticthat he did not want to know if he had DKC. During testing, it wasdiscovered that he had unusually short telomeres and would mostlikely develop symptoms of DKC. Why might mutations in genes encoding telomerase subunitslead to bone marrow failure?