7. Consider the following plasmid (size 6700 bp), with restriction sites at the positions indicated: BamHI + 1 6700 bp 2800 3500 EcoRI BamHI Probe c) This gel is transferred to a membrane in a Southern blot experiment, and hybridised to a radioactively labelled 100 bp probe, which anneals to the plasmid DNA at the position indicated on the diagram above. Draw the autoradiograph profile you would expect to observe for the membrane, indicate the sizes of the bands on the blot.

7. Consider the following plasmid (size 6700 bp), with restriction sites at the positions indicated: BamHI + 1 6700 bp 2800 3500 EcoRI BamHI Probe c) This gel is transferred to a membrane in a Southern blot experiment, and hybridised to a radioactively labelled 100 bp probe, which anneals to the plasmid DNA at the position indicated on the diagram above. Draw the autoradiograph profile you would expect to observe for the membrane, indicate the sizes of the bands on the blot.

Biology: The Unity and Diversity of Life (MindTap Course List)

15th Edition

ISBN:9781337408332

Author:Cecie Starr, Ralph Taggart, Christine Evers, Lisa Starr

Publisher:Cecie Starr, Ralph Taggart, Christine Evers, Lisa Starr

Chapter15: Studying And Manipulating Genomes

Section: Chapter Questions

Problem 2SQ

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Match the method with the appropriate enzyme. _____ PCR a. Taq polymerase _____ cutting DNA b. DNA ligase _____ cDNA synthesis c. reverse transcriptase _____ DNA sequencing d. restriction enzyme _____ pasting DNA e. DNA polymerase (not Taq)
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1. Briefly explain why the total size of the pMBBS plasmid in the Restriction Enzymes practical is 3000bp (base pairs) 2. Briefly explain why the cut sites on the pMBBS plasmid in each Restriction Enzymes (EcoRI, BamHI, and XhoI) just 1 each 3. Briefly explain what led to the 5 fragments formed which linked to the 500bp, 1000bp, 1500bp, 2000bp, 2500bp sizes
1 (a) What do ApR, TcR, and ori on the pBR322 map represent and discuss there individual functions? (b)Does the undigested plasmid show more than a single band when electrophoresed? Why? (c)What other kinds of molecules, in addition to plasmid DNA would you expect to find in a sample of plasmid DNA extraction? (This is not a graded assignment)
1. Write if the statement is true and modify the statement with the correct answer if its false a.White colonies represent those bacteria that do not express the genes for beta-galactosidase because restriction enzymes have cut the gene and ligase have not repaired the break b.Antibiotic resistance genes in bacterial transformation are used as marker genes because of ease of detection of transformants. c. Bacterial transformation simply means the insertion of a recombinant plasmid into a bacterial cell.
1. Match the features of plasmids below with their meaning : Origin of replication LacZ gene antibiotic resistance gene Multiple cloning site A.required for maintenance of the plasmid in E. coli cells used to select for coli that contain our plasmid B. Required for replication of plasmid DNA inside an E. coli cell C. An engineered region of the plasmid that contains many unique restriction enzyme recognition Sites D. Allows for screening for plasmids with an insert in the multiple cloning site
1. Given the following restriction endonucleases and the sequences of their corresponding restriction sites, which would be LEAST useful for cutting the plasmid and the foreign DNA to be inserted? [The arrows indicate where cuts are made.] (see img) a. EcoRI b. HindIII c. HaeIII d. BamHI 2. EcoRI and HindIII are two different restriction enzymes. If the DNA from two different sources were cut with either EcoRI or HindIII, which of the cut DNA fragments would NOT join together easily so that they could be sealed with ligase? a. E. coli DNA cut with EcoRI and human DNA cut with HindIII b. E. coli DNA cut with HindIII and mouse DNA cut with HindIII c. human DNA cut with EcoRI and chimpanzee DNA cut with EcoRI d. mouse liver DNA cut with HindIII and mouse kidney DNA cut with HindIII
1. You have the plasmid pUC18/19, which is a circular plasmid that consists of 2686 bp. What would the number of and length of the fragments be if you cut the plasmid with the following restriction enzymes or combination of enzymes? Give a schematic representation of the digestions.a. PscI & GsuIb. ScaI, PdmI & BsaXI c.ScaI, SspI & EheI
3A. Following transformation with the CRISPR plasmid, the yeast will be plated on SD-URA plates. Why is SD-URA media used? Name the relevant marker gene in the pCRCT CRISPR plasmid. 3B. What would you expect to see if you plated on YED accidentally? Will most of the yeast be red or white? Why? 3C. The complete genotype of the host yeast strain is MATa ade2 his3 leu2 met15 ura3. Name one advantage of using a yeast strain with auxotrophies for several genes (i.e. his3, leu2 and ura3). 3D. After successful CRIPSR editing, the yeast can later be “cured” of the recombinant CRISPR plasmid – that is, the plasmid is lost but the CRISPR edit is stably inherited. Write the new genotype of the yeast-based on the ADE6 gene.
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6. Which of the following steps are catalyzed by Taq DNA polymerase in a PCR reaction?a. Denaturation of template DNAb. Annealing of primers to template DNAc. Extension of primer end on the template DNAd. All of the above Please also explain the answer properly and refer to NCERT XII (chapter 11) textbook.
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