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MCDB 101A M 2013 2nd Midterm Name_____________ _____________________________

7/22/13 (100 points total) Perm #__________________________________________









Do not open exam until instructed to do so

Write your name, perm#, & MCDB 101A M2013 Midterm 2 on your Scantron Form






































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Answer questions 1-11 on a Scantron form (3 points for each correct answer). For each multiple choice
question mark the letter corresponding to the best answer. For the True/False question mark a for true and
b for false.

1. Which is true of RecA protein?
a. It cleaves CII protein and as a result prevents the establishment of the lysogenic state
b. It is required for excision of lambda from its host DNA in response to DNA damage
c. It is cleaved by LexA in the SOS response pathway
d. It plays a role in the repair of thymidine dimers
e. Both a and d

2. Which of the following genotypes would result in the production of -galactosidase but not permease in
the absence of inducer? (for this problem assume mutations in lac operon genes are nonsense mutations)
a. I
S
P+O
C
Z+Y / I +P+O+Z + Y+
b. I P+O+Z+Y+ / I
- d
P O+Z - Y+
c. I P+O+ Z+Y / I+P+O+Z Y+
d. I
S
P O+Z+Y+/ I + P+O
C
Z Y+
e. both a and c

3. Which is true of a silent mutation?
a. It has no phenotype
b. It is located in the promoter region of a gene
c. It changes an amino acid to another of similar size and biochemical characteristics
d. It produces a wild type phenotype
e. It can result from insertion of a single nucleotide into the protein coding sequence of a gene

4. You run a gel retardation assay on a lac I protein isolated from E. coli. No shifted band is observed when
you combine the protein with radioactively labeled lacO DNA (4 points). This result is consistent with which
form(s) of the lac repressor?
a. Lac I -
b. Lac I
S

c. Lac I
-d

d. a and c
e. a, b, and c

5. A mutation in the gene encoding which of the following proteins would enhance the sensitivity of the Ames
test to the type of mutations produced by nitrites?
a. Uracil glycosylase
b. Catalase
c. Vsr
d. MutS
e. Both a and c

6. Shown to the right is a base pair that resulted from tautomerization. The bases are
a. enol cytosine and imino adenine
b. keto guanine and enol adenine
c. imino thymine and enol cytosine
d. keto thymine and enol guanine
e. pseudoura cil and amino xanthine


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7. Which correctly describes the blue-white cloning system?
a. White colonies indicate successful integration of your gene into the cloning plasmid
b. A multi-cloning site is located upstream of the lacZ gene contained in the cloning plasmid
c. The N-terminal portion of the lacI gene is present in the cloning plasmid
d. A dominant mutation is present in the plasmid-encoded copy of lacI
e. A wild type copy of lacZ is present in the genome

8. Which of the following is used in PCR?
a. A polymerase that can survive repeated cycles of freezing and thawing
b. The deoxy nucleotides dATP, dUTP, dCTP, and dGTP
c. A pair of RNA primers
d. DNA ligase
e. None of the above

9. Phage lambda immunity is a direct consequence of the action of which of these proteins?
a. N
b. CI
c. CII
d. Cro
e. Q

10. Which of the following would have been an appropriate conclusion if samples taken from the bulk culture
in Luria and Delbrooks fluctuation experiment exhibited a greater variation in colony number than was
observed among samples taken from the individual cultures?
a. Mutations were induced by exposure of the bacteria to phage
b. Random mutations occurred prior to exposure of the bacteria to phage
c. Early mutations were random; late mutations were induced
d. The mutation rate was greater than the mutation frequency
e. No conclusions on the mechanism of mutagenesis could be made from this experiment

11. True or false. Depletion of nutrients required for the growth of a bacterium that contains a lambda
prophage will tend to maintain the phage in a lysogenic state. (3 points).

12. The binding of CI to the O
R2
operator site is facilitated by CI binding to the O
R1
site by a process known as

_________ ______________________________________ binding. (3 points)

13. (12 points) Shown below are genotypes of the lac operon and its regulatory genes. C is the wild type
allele of the CAP protein, C

is a recessive allele that produces a protein that cannot bind to the CAP site &
C* is a dominant negative allele that produces a CAP protein that cannot bind cAMP. Fill in the table with
either +or 0 to indicate whether -galactosidase is produced under the indicated conditions.

-galactosidase
Genotype -lac
+glu
-lac
-glu
+lac
+glu
+lac
-glu
C
+
I

P
+
O
+
Z
+
Y
-
/C
+
I
+
P
+
O
C
Z

Y
+

C
+
I
+
P
+
O
+
Z
+
Y
+
/C

I
-d
P
+
O
+
Z
-
Y
+

C* I
S
P
-
O
+
Z
+
Y
-
/C
+
I
+
P
+
O
C
Z
+
Y
+



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14. (6 points) One way in which a western blot differs from a Southern blot is the method of transferring the
target material from the gel to a nitrocellulose membrane. Give a one sentence description of how this
transfer is accomplished for each of the two blotting methods, indicating in your answer the biochemical
nature of the target material for each assay e.g lipids, carbohydrates etc.

Southern blot:




Western blot:




15. The sequence AUGCAUGGUAAC is found within a mRNA molecule. Bacteria containing this mRNA
were treated with proflavin, resulting in a single point mutation within the corresponding DNA sequence.
The amino acids translated from the mutated sequence have the sequence: Met-His-Trp.

a. (4 points) Write out the DNA sequence of the template (non-coding) strand as it appears after the
mutation has occurred. Label the 5 and 3 ends of your sequence.




b. (3 points) Is this mutation a transition, transversion, deletion, or insertion? (circle correct answer,)


16. (6 points) Shown below are three pairs of PCR primers that are complementary to DNA sequences
upstream and downstream of a gene you wish to amplify. Two of these primer pairs would not be good
choices for use in a PCR reaction. List the two sets of primers that should not be used and give the reason
why each would be unsuitable for PCR.

Set #1: 5ACTGCCTACTTGAGATGC 3 5GCAACTGTCAGGTACATG 3

Set#2: 5 CTCATACGAACTGAGCTA 3 5TCCGATACTATGATTCAA 3

Set#3: 5 CGATCGATACTATGACCG 3 5CGGTCATAGTATCTGATC 3








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17. (5 points) Explain why a small amount of histidine is added to growth media used in the Ames test.









18. (5 points) Experiments were carried to study the ability of uvrA deletion strains of E. coli to survive UV
irradiation. Wild type and mutant strains were spread on solid media, the plates were exposed to UV light,
and colonies were counted after 2 days. One mutant produced more colonies than the wild type strain. You
learn that all of the strains except for the highly resistant mutant were incubated in a drawer. The latter was
incubated on the bench top because there was insufficient room in the drawer to contain all of the plates. Give
an explanation that does not involve the uvrA gene for the UV resistance of the mutant strain.














19 a. (4 points) What potential problem do tRNA suppressor mutations create for wild type protein-coding
genes in a genome?







19 b. (3 points) Give one reason why the problem you described in 19 a. turns out not to be a significant issue
for the cell.







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20. You grow 10 cultures of T1 sensitive E. coli to a density of 1 X 10
9
bacteria/ml. You take 0.1 ml of each
culture and add it to a dish of T1 bacteriophage. You obtain the following #s of colonies:

dish 1 2 3 4 5 6 7 8 9 10
#colonies 0 12 5 0 0 0 75 0 0 8
Write out the appropriate formulas and show all work in your answers to parts a and b.
a) (6 points) Calculate the rate of reversion of T1 sensitive bacteria to a T1 resistant phenotype







b) (5 points) Calculate the frequency of reversion to T1 sensitivity








21 a. (2 points) With what repair process is DNA adenine methylase associated?





21 b. (3 points) Would a mutation in the DNA adenine methylase promoter resulting in increased production
of DNA adenine methylase be likely to lead to an increased or decreased level of DNA repair as compared to
the normal level of expression of this gene?