Microbial Genetics Exam          Fall 1999

Your Name:____________________                                     

1.      (10 points) Explain in molecular terms why, when l_B is used to infect E. coli K12, the E.O.P. is 10^-4 and not one or zero.

2.      (12 points) Describe the hok/sok system of partition control of plasmids.  Would a wild-type bacterium be able to maintain a plasmid with a hok⁺/sok^– genotype?  Why or why not?

3.      (10 points) Compare and constrast Type I and Type II restriction/modification enzymes.

4.      (10 points) What is the lacZa (lacZ') fragment, which is often found on cloning vectors?  What is role of the multiple cloning site (or polylinker) located in the lacZa fragment?

5.      (10 points) Why are 6 base cutting restriction endonucleases usually not suitable for generating 1 kb ("gene sized") fragments?  Can 4 base cutters generate 1 kb fragments, and if so, how

6.      (12 points) What is a Northern blot experiment?  What types of information can it tell you about the transcription of a gene?

7.      (12 points) Describe the important features of cloning vectors.  Explain the purpose of the antibiotic resistance gene(s) found in typical plasmid vectors. 

8.      (12 points) Describe the features of the E. coli expression system that involves the T7 RNA polymerase — the pET system.  What is the function of the lacUV5 promoter in this system?

9.      (12 points) A circular DNA molecule is digested with restriction endonucleases, and the following results are obtained: EcoRI, 12 kb; PstI, 9 kb, 3 kb; HindIII, 12 kb; EcoR1/PstI, 9, 2, and 1 kb; EcoRI/HindIII, 11 and 1 kb; PstI/HindIII, 9 kb and 3 kb.  Given the EcoR1 and HindIII sites below, map the PstI site(s).