ps7s - MIT Department of Biology 7.014 Introductory...

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MIT Department of Biology 7.014 Introductory Biology, Spring 2004 7.014 Spring 2004 PS 7 answers 1 of 7 Solutions to 7.014 Problem Set 7 Question 1 On your trip to the Amazon you were introduced to a new plant that the indigenous people use as an anti-viral treatment. You took samples home to your lab and you found that this plant makes a protein (the PV protein) that prevents viral replication. Excited by the possible anti-AIDS applications, you construct a genomic DNA library from this plant in the hope of cloning the PV gene. a) What is a genomic DNA library? A collection of bacterial cells (or phage particles) each of which contain a different small piece of the genomic DNA that you are interested in. b) You get a DNA sample from cells, digest it with a restriction enzyme, and clone it into a vector. List 3 features of the vector that are absolutely required for your library construction. An origin of replication (ori) A cloning site A selectable marker c) Circle on the following lists ALL you would need in order to construct the genomic DNA library. Assume you start with intact plant genomic DNA. Restriction enzyme Ligase Enzymes Reagents Size separation gel Okasaki fragments DNA Polymerase ATP, TTP, CTP, GTP RNA Polymerase ddATP, ddTTP, ddCTP, ddGTP Transcriptase Primers Reverse Transcriptase 3' to 5' exonuclease Cloning vector Replication fork E. coli (bacteria) Human cells Virus Briefly describe the function of each item circled. Restriction enzyme: cut the vetor and the genomic DNA Ligase: join together the cut DNA fragments Cloning vector: receives the cut genomic DNA and allows propagation in the bacterial cell host. E. coli: Acts as a host that receives the recombiant plasmids and replicated this new DNA 1
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7.014 Spring 2004 PS 7 answers 2 of 7 Question 1, continued d) Your genomic DNA library is now ready. You have PV protein sequence information and want to make a DNA probe that is 12 nucleotides long. Given the protein sequence below, circle the amino acids that you will use to design your DNA probe. Why did you choose these amino acids? leu-arg-met-gly-ser-ala-val-leu-pro-thr-arg-ser-trp-met-tyr-lys-arg-his-thr-ser-ala-cys-met-pro-phe These amino acids can be encode by fewer possible DNA sequences that any other four consecutive amino acids.
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ps7s - MIT Department of Biology 7.014 Introductory...

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