BCHM Exam 3 Study Guide - BCHM Exam 3 Study Guide...

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BCHM Exam 3 Study Guide Polymerase Chain Reaction - making multiple copies of the gene of interest 1. Components a. template DNA strand b. dNTPs c. DNA polymerase i. Taq – thermophile so it won’t denature at high temperatures d. buffer & ATP e. DNA primers 2. 3 steps a. denature: heat at greater than 9 C instead of using helicase 0 0 b. anneal: cool at 4 C – binds primers & anneals the strands 0 0 c. primer extension: DNA polymerase does its job close to 7 C 0 0 3. 2 n gives you the # of DNA molecules, where n = # of PCR cycles 4. primers only bind to 3’ end so they can be built 5’ to 3’ – otherwise PCR would run in the reverse direction 5. be able to identify the correct primer pair Transcription 1. one gene one enzyme hypothesis a. isolate mutant that cannot grow arginine and grow in 3 different mediums – test to see if enzymes required for different steps in the pathway for synthesizing arginine are missing i. ornithine supplement can only grow arg 1 mutant ii. citrulline supplement can grow arg 1 and arg 2 mutants iii. arginine supplement can grow all 3 arg mutants b. even if the gene itself is flawed, if the medium is supplemented with what the gene SHOULD make, the mutants should grow c.
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