Name:_______________________

BioSc1940 Molecular Biology

Midterm Exam 1
September 29, 1999
Instructors: Hendrix and Arndt

Multiple choice (2 pts each)—Circle one answer for each

1. According to the "Central Dogma", all of the following transfers of

information happen except:

A. RNA to DNA
B. DNA to DNA
C. protein to protein
D. RNA to protein
E. DNA to RNA

2. Thirty-five years ago, in the psychedelic '60's, hippies ruled the world and
there was a march on Washington DC every few months to protest the war in
Vietnam. In molecular biology, all of the following were true except:

A. restriction enzymes were under study

B. the double helical structure of DNA was known
C. DNA cloning had re-invigorated the study of Drosophila
D. mRNA had been discovered and its role in gene regulation was under
intense study
E. none of the above

3. The restriction enzyme SalI cuts the sequence GTCGAC to leave a four base,
5' overhang. The restriction enzyme XhoI cuts the sequence CTCGAG to
leave a four base, 5' overhang. If you mix a SalI-digested insert with a XhoI-
digested plasmid vector and perform a ligation, which best describes the
properties of your ligation product:

A. The insert can be removed by digesting with SalI or XhoI.

B. The insert can be removed by digesting with SalI but not XhoI.
C. The insert can be removed by digesting with XhoI but not SalI.
D. The insert cannot be removed by digesting with either SalI or XhoI.
E. You have no product because the insert and vector cannot be ligated
together.

1
 Name:_______________________

4. In the Meselson-Stahl experiment that demonstrated semi-conservative

replication of DNA, the most critical piece of evidence supporting the
conclusion was:

A. replicated DNA appeared that had fully light density

B. DNA could not replicate due to the unnaturally high concentration of heavy
isotopes
C. the distribution of DNA among the different density classes changed with
time
D. DNA of fully heavy density did not survive the first round of replication
E. the mass ratios among the different density classes continued to change past
the first round of replication

5. Restriction enzymes are found in virtually all bacterial species, and their
sequence specificities cover almost all possible short (4-6 bp) sequences. Why
do you think this is the case?

A. Restriction enzymes were put on Earth by a benevolent deity to make the

lives of molecular biologists easier
B. Restriction enzymes provide protection for the cell against incursion of
foreign DNA
C. Restriction enzymes provide the means by which bacteria carry out gene
cloning
D. Restriction enzymes provide a means to ensure that the cells’ DNA is
properly and specifically methylated
E. Restriction enzymes recognize as foreign any DNA that contains copies of
their recognition sequence and destroy it

6. You are studying a new restriction enzyme that recognizes and cuts at the
sequence 5’-GCGCNNNNNGCGC-3’. How big on average do you expect the
restriction fragments to be when you digest the genomic DNA of the hairy
aardvark?

A. ~250 bp
B. ~1 kb
C. ~4 kb
D. ~16 kb
E. ~64 kb

2
 Name:_______________________

7. When a typical restriction enzyme like BamHI cuts DNA, it leaves:

A. a 5' PO4 on the top strand and a 3' PO4 on the bottom strand
B. a 3' PO4 on the top strand and a 5' PO4 on the bottom strand
C. 5' PO4's on both strands
D. 3' PO4's on both strands
E. the PO4 at the site of each cut is deleted and replaced later by DNA ligase

8. Which of the following is not generally used for site-directed mutagenesis of

DNA:

A. a mutagenic oligonucleotide
B. dimethyl sulfate
C. a dut- ung- strain of E. coli
D. DNA polymerase
E. plasmid vectors derived from M13

9. Using a standard PCR reaction, you can:

A. invert a cloned piece of DNA in a plasmid vector

B. insure your survival of everything except falling redwood trees
C. determine unknown DNA sequence
D. synthesize short oligonucleotide primers
E. amplify DNA between known sequences

10. DNaseI footprinting experiments:

A. require a DNA probe labeled at one end of one strand

B. reveal the sequence to which a protein binds
C. give quantitative information about the affinity of a protein for the DNA
D. require use of a denaturing polyacrylamide gel
E. all of the above

3
 Name:_______________________

True or false (2pts each) —indicate your answer clearly

11. Accurate mapping of the 3' end of an RNA transcript can be performed either
by primer extension experiments or S1 nuclease assays.

12. Electrophoretic mobility shift assays can be used to monitor protein complex
assembly on a DNA fragment but do not reveal the actual sequences
recognized by the proteins.

13. Sigma (σ) factor has an important role in transcription termination at factor-
dependent termination sites.

14. Once RNA polymerase has made the first phosphodiester bond, it’s like a
hound on the scent and doesn’t ever stop until it reaches the termination
signal at the end of the mRNA sequence.

15. The high resolution structure of bacterial RNA polymerase strongly suggests
that the DNA double helix in the elongation complex bends more than 90° as
it passes through the RNA polymerase.

16. RNA polymerase determines which nucleotide to put in each position of the
RNA molecule by reaching into the major groove of the template DNA and
contacting the sides of the bases.

4
 Name:_______________________

Short answer questions (6pts each)

17. Shown below is an autoradiograph of a DNA sequencing experiment performed by

the method of Sanger. What is the sequence of the DNA? Indicate polarity.
G A T C

You perform Sanger and Maxam-Gilbert sequencing reactions to show the positions of
guanines in your DNA sample. If you run the products of these reactions side-by-side
on the same denaturing polyacrylamide gel, will a Sanger-generated band
corresponding to any given G in your DNA sequence run with the same or a different
mobility in the Maxam-Gilbert lane? Briefly explain your answer.

18. Shown below is a restriction map for the beta-globin gene from mouse. You
digest mouse genomic DNA with EcoRI and resolve the digestion products on
an agarose gel.

1.0 kb 1.8 kb 0.6 kb

beta-globin

probe

What do you expect to see if you stain the agarose gel with ethidium bromide?

What do you expect to see if you perform a Southern blot on the agarose gel
using the radiolabelled DNA probe indicated on the figure (hatched box)?

5
 Name:_______________________

19. You have just completed a C0t analysis of the genomic DNA of the hairy
aardvark. Your technician brings you a plot of the data:

Oops! Your technician forgot to label the axes of the graph.

a) Please add the correct labels.

b) State (briefly) what this graph allows you to conclude about the hairy
aardvark genome.

20. Prokaryotic transcription terminators of the factor-independent type rarely

have the same sequence as each other, but they virtually always have the same
sequence characteristics. Write out below a DNA sequence that you believe will
convince RNA polymerase to stop transcribing and release its RNA transcript.
Show the sequence of both strands, indicate 5' and 3' ends, and indicate the
direction of transcription.

6
 Name:_______________________

7
 Name:_______________________

21. Which subunit of bacterial RNA polymerase has the catalytic activity?

Which has the primary DNA binding activity?

22. You have a library of genomic DNA of the hairy aardvark cloned in
a λ vector. You wish to clone and study the gene for isocitrate dehydrogenase.
Your friend who studies the piebald peckary has cloned the piebald peckary
isocitrate dehydrogenase gene and has sent her clone to you. Outline very
briefly how you will proceed from here.

8
 Name:_______________________

Long answer (16 pts each)

23. Ribonucleoside triphosphates are polymerized by RNA polymerase into

RNA. 2’-deoxyribonucleoside triphosphates are polymerized by DNA
polymerase into DNA. What do you think would happen if you added a small
amount of a 3’-deoxynucleoside triphosphate (say 3’-deoxy ATP) to each of those
two reactions? Can you use this information to design a new method for
determining DNA sequence? What else might your method be useful for?

9
 Name:_______________________

24. Consider the UP element found in some particularly strong prokaryotic

promoters. Diagram such a promoter, showing where the various promoter
elements (including the UP element) are found. Does RNA polymerase interact
directly with the UP element? If so, what part of RNA polymerase? Describe in
moderate detail the nature of the experiments that were used to answer these
questions, including the results of the experiments.

10