ProblemSet_01_Solutions

ProblemSet_01_Solutions - MCB 121 Spring 2010 Problem Set 1...

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1 MCB 121 Spring 2010 Problem Set 1 You are studying a new bacteriophage you have discovered named T100. This phage is morphologically similar to T4 and you have determined that it undergoes a similar process of replication and assembly during infection in E. coli . You have isolated three temperature sensitive mutants of T100 and have determined that these contain mutations in separate genes. In addition, you have used electron microscopy to analyze the viral products produced following infection of E. coli at the non-permissive temperature of 42°C (see table). Finally, you have determined that E. coli extracts prepared following infection with each mutant at 42°C can complement extracts prepared from the other mutants to produce active virus. Mutant Gene Viral Structures Produced Inferred Defect 1 Complete heads and tail fibers Tail formation 2 Complete tails and tail fibers Head formation 3 Heads and tails and tail fibers but head and tail remain unattached ? 1. You entertain three possible functions for Gene 3: (1) involvement in head maturation, (2) tail maturation, (3) or head-tail joining. Design an experiment to distinguish between these possibilities and explain what results you would expect for each. For your experiment, assume that you can purify complete tails and heads from mutants 1 and 2, respectively. Answer : One approach would be to test whether an extract made from E. coli cells infected with Mutant 3 at the non-permissive temperature (we will call this a “Mutant 3 Extract”) can form complete and infectious viral particles when combined with purified heads or purified tails (obtained from mutants 1 and 2, respectively). The expected outcomes would be: Possibility 1: Gene product 3 is involved in head maturation. In this case, the Mutant 3 extract will contain heads that are not complete but the tails and tail fibers should be fine. Therefore, functional virus should be produced when this extract is mixed with purified complete heads but not purified tails. Possibility 2: Gene product 3 is involved in tail maturation. Following the same reasoning as above, in this case the Mutant 3 extract should contain defective tails but normal heads and tail fibers. Functional virus should result when this extract is mixed with purified complete tails but not purified heads. Possibility 3: Gene product 3 is involved in the joining of heads and tails. In this case, because purified heads or tails are being added to an extract that is defective in joining heads and tails, no functional virus will be produced in either case. Note that this is constitutes a “negative result” and thus it would be impossible to conclude based upon this result alone that Gene 3 is involved in head-tail joining. The results would simply be consistent with such a role. Other
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2 experiments would have to performed, ultimately purifying this factor following infection with wild type virus and demonstrating that the gene encoding this protein is indeed Gene 3. 2.
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ProblemSet_01_Solutions - MCB 121 Spring 2010 Problem Set 1...

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