Midterm2'09Key - R. L. Rodriguez, Instructor BIS101-001,...


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R. L. Rodriguez, Instructor BIS101-001, Spring 2009, Midterm 2 May 13, 2009 Please print your name below and each subsequent page. Your exam has 9 questions and 8 pages (including cover page and score sheet). 1. In prokaryotes, several enzymes are responsible for replicating chromosomal DNA. The table below shows enzymatic activities for these enzymes. Using plus (+) and minus (-), indicate which enzymes are associated with which activities. (7 pts.) Activities Pol I Primase Pol III Ligase 5’-3’ exonuclease X 3’-5’ exonuclease X X 5’-3’ deoxynucleotide synthesis X X ATP-dependent 3’-5’ joining X 5’-3’ ribonucleotide synthesis X E. coli DNA polymerase makes approximately 9 errors per million bases synthesized while Taq DNA polymerase makes 285 errors per million bases synthesis. Which of the above enzymatic activities is responsible for this difference? (3 pts.) 3’ to 5’ Exonuclease 2. As can be seen in the steps below, the polymerase chain reaction (PCR) involves the cycling of DNA and primers between different temperatures (in degrees Celsius). In the blanks below, fill in the temperature best suited for a particular step. (8 pts.) (+, - 2°C) Step 1: Melting. The mixture is heated at __ 96°C ____for 5 minutes to ensure that the DNA strands as well as the primers have melted. The DNA-Polymerase can be present at initialization, or it can be added after this step. Step 2: Annealing. By reducing temperature to __ 68°C ___ for 30 seconds primers can anneal to target sequence in single stranded DNA. Step 3: Elongation. If DNA polymerase is present, deoxynucleotides can be elongation at __ 72°C __ for 45 seconds. Step 4: Termination. Mixture is held at _ 4°C _ until it can be analyzed by gel electrophoresis.
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Name____________________________________________________ 2 3 . In the PCR reaction, temperature is used to denature the double strands of target sequence DNA and anneal the oligonucleotide primers. Temperature is not used to denature DNA in Southern blotting. Based on your knowledge of the forces and interactions that stabilized the
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