MOL

214
Exam 1

October 22, 2015




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 Exam Code Number: ______________

Multiple Choice (2 points each)

1. Which of the following correctly describes the order of events in DNA replication?

1. Okazaki fragments are joined together

2. DNA is synthesized by polymerase
3. Primase synthesizes primers
4. RNA primers are removed by exonucleases
5. Helicase unwinds DNA
6. Single-stranded binding protein binds DNA
7. Gaps are filled by polymerase

a. 5, 6, 3, 2, 4, 7, 1
b. 5, 2, 3, 6, 1, 7, 4
c. 3, 6, 5, 2, 4, 1, 7
d. 3, 5, 6, 2, 1, 7, 4

2. Which of the following is involved in transcription termination?

a. Sigma factor
b. Promoter
c. Stem-loop sequences in mRNA
d. RNA polymerase encountering another RNA polymerase
e. Ligase

3. Before Watson and Crick solved the structure of DNA, Linus Pauling, a chemist at Caltech,
proposed that DNA was a triple helix. Based on what we know now, which of the following
properties of DNA conflicts the most with Pauling’s model?
a. DNA is soluble in water
b. Chargaff’s base ratios (%A = %T and %G = %C)
c. DNA strands are anti-parallel to each other
d. DNA phosphodiester bonds involve a single phosphate

4. Which of the following features is common to both DNA replication and transcription?
a. Nucleotides are added to the 3' end of the newly synthesized strand
b. A sugar-phosphate bond is formed between the 3' hydroxyl and the 5' phosphate
c. Deoxyribonucleotides are incorporated into the growing sequence
d. Both RNA and DNA polymerase require oligonucleotide priming
e. Both RNA and DNA polymerase initiate at promoter sequences

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5. DNA polymerase builds DNA in the following direction:

a. 5’à3’ in the leading strand and 3’à5’ in the lagging strand
b. 3’à5’ in the leading strand and 5’à3’ in the lagging strand
c. 5’à3’ on both strands
d. 3’à5’ on both strands

6. Which of the following mutations is characteristically caused by UV damage?

a. Thymidine dimers
b. Deamination
c. Depurination
d. Nitrosamines

7. The cell membrane can be described most accurately as

a. permeable to most small molecules but impermeable to larger ones.
b. permeable to only larger molecules.
c. permeable to all molecules.
d. permeable to some molecules and impermeable to others.
e. impermeable to all polar molecules.

8. Heating DNA to 98°C breaks which kind of bonds?

a. Covalent bonds
b. Ionic bonds
c. Phosphodiester bonds
d. Hydrogen bonds
e. It does not break any bonds

9. Which of the following statements about sigma factors is true?

a. Sigma factors are important for the initiation of transcription in both eukaryotes and
prokaryotes
b. Sigma factors bind to the promoter and recruits RNA polymerase to initiate transcription.
c. Sigma factors bind to RNA
d. Sigma factors are important for the termination of transcription in both eukaryotes and
prokaryotes.

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10. Examine the graph shown below. Which of the lines shown on the graph below would be
characteristic of a mutant lacking telomerase?

a. Line A
b. Line B
c. Line C
d. Line D

11. During replication of the template strand shown below, DNA polymerase adds the wrong base to
the newly replicated strand generating a mismatch. Which of the choices below represents the
correct way to repair the mismatch?

Template Strand
T
C New Strand

a. c.

T T

A G

b. d.

G A

C C

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Short Answer Questions:

12. Label which arrows on the replication bubble shown below are leading and lagging.
On the diagram indicate where topoisomerase would be located. (3 points)

topoisomerase topoisomerase
leading lagging

lagging
leading

No credit given if topoisomerase is drawn at the fork

13. How would Avery, McCarty, and MacLeod’s conclusion be different if protease had destroyed
the transforming principle? (2 points)

Such a result would suggest that protein was transforming principle and that it was the genetic
material.

Follow this scenario to the Hershey-Chase experiment: which radioactive isotope would have
entered bacteria upon infection with phage? (1 point)
35
S

14. Microsatellites (also known as short tandem repeats or STRs) are a powerful forensics tool used
by law enforcement. Why is this the case? (3 points)

There is a high degree of variability/polymorphism at STRs . (2 points)

Combining 13 STRs ensures that no two people will have the same set of alleles allowing
identification of an individual. (1 point)

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15. You are given two samples of DNA, one from Clostridium perfringens and the other from
Mycobacterium tuberculosis. Unfortunately, the labels on the tubes of DNA were accidently
removed, and you do not know which DNA sample belongs to which organism. As you were
doing some work with Escherichia coli in the lab, you also have some of its DNA available as
well. The % G+C values for each of the organisms is known (C. perfringens, 27%; M.
tuberculosis, 67%; E. coli, 50%). Based upon this information label the melting curve for each
organism on the graph below. (3 points)

A
B

C

A is C. perfringens, 27%; B is E. coli, 50% and C is M. tuberculosis, 67%

16. Telomerase is a protein with an RNA component. What is the role of the RNA component? (2
points)

The RNA component of telomerase provides a template for synthesis of the telomere repeats.

Would telomerase still be able to perform its function without the RNA component? Why or why
not? (2 points)

No, telomerase would not be able to function without the RNA component because then would
be nothing to serve as the template for the telomeres.

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17. Meselson and Stahl demonstrated that DNA replication was semi-conservative by allowing cells
to replicate in media containing 15N, then moving them to media containing 14N and subjecting
the DNA to equilibrium density centrifugation. DNA from the starting cells (grown in 15N)
would look like the diagram below after density centrifugation:

Less Dense 14
N 15
N More Dense

Draw what the results would look like after 1, 2 and 3 rounds of replication if DNA replication
was conservative. (6 points)

One round of replication

Less Dense 14
N 15
N More Dense

Two rounds of replication

Less Dense 14
N 15
N More Dense

Three rounds of replication

Less Dense 14
N 15
N More Dense

18. You are working in your senior thesis lab and want to amplify the sequence depicted below.
Design primers 9 base pairs in length written 5’à3’ to amplify the entire sequence. (4 points)

5’- ATGCGCTTGATTCGCATGCGCGAATTACGTCGACTC-3’
3’- TACGCGAACTAAGCGTACGCGCTTAATGCAGCTGAG-5’

Forward primer: 5’-ATGCGCTTG-3’

Reverse primer: 5’-GAGTCGACG-3’

Which of your primers would have the higher melting temperature? Why? (2 points)

The reverse primer would have a higher melting temperature as it has a higher GC content

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19. Scientists discovered a new nucleotide polymerase that does not need a template or a primer, and
only uses purines. The product of the polymerase is shown below:

Which of the following nucleotides would act as a terminator for this polymerase? Circle all that
apply. (4 points)

The phosphodiester bonds are between the 2’ and 5’ hydroxyls. A and B both lack 2’-OH and
will act as chain terminator.

20. Why is topoisomerase not required for the replication of DNA during PCR? (2 points)

Because heat is used to separate the DNA there is no problem with supercoiling of the helix.

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21. Restriction enzymes are useful for determining the orientation of inserts cloned into plasmids.
Given the following information, draw a restriction map of this hypothetical 8 kb plasmid. Please
note that there may be more than one acceptable way to draw the map, but you only need to
show one.
On the map indicate the location of each of the following restriction enzymes, and show the
length (in kb) between the restriction enzyme sites (6 points)
• This plasmid can be cut by BamHI, XbaI, PstI and HindIII.
• Each of these cuts only once
• Restriction digestion followed by DNA gel electrophoresis yields bands of the
following sizes:

HindII+ XbaI=1kb and 7kb
BamHI +XbaI= 3kb and 5kb
BamHI+ HindII= 4kb
HindII + PstI= 3kb and 5 kb
PstI+ XbaI= 2kb and 6kb

HindIII
1
XbaI

4 8kb 2

PstII
1
BamHI

If this hypothetical plasmid were a cloning vector, what three features would it need to contain
and why? (3 points)
Origin of replication – needed to ensure plasmid is replicated.
Selectable marker – allows for selection of cells that contain the plasmid
Cloning sites – restriction enzymes used to insert DNA into vectors.
(Also accepted lacZ as a way to tell that a recombinant plasmid containing an insert was created)

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22. You have been studying eukaryotic RNA polymerase II and have discovered a mutant version of
the enzyme that cannot be phosphorylated in its C- terminal domain (CTD). However, you have
also discovered Drug X that allows RNA polymerase II to begin transcription without CTD
phosphorylation. What will happen to mRNAs transcribed by this mutant polymerase that has
been treated by Drug X? (4 points)

The mRNAs will not be processed properly. Although transcription occurred because of Drug
X, the CTD still lacks phosphate groups and cannot recruit capping enzymes, splicing
machinery, or the cleavage and polyadenylation factors. Therefore, the mRNA will not mature
and should not be exported to the cytoplasm.

23. Examine the promoter shown below and answer the following questions.

-35 -10 +1

5’ ATGGAGCGCTATCGCATGA 3’
TTGACA TATAAT

3’ TACCTCGCGATAGCGTACT 5’
AACTGT ATATTA

What type of organism would you find a promoter like the one shown above? (2 points)
This type of promoter is found in bacteria

What factors will bind and recognize this DNA sequence? (2 points)

Sigma factors recognize the bacterial promoter and recruit RNA polymerase to begin
transcription.

What will the sequence of the RNA transcript be for the gene shown above? (please write it
5’à3’) (3 points)

5’ AUGGAGCGCUAUCGCAUGA 3’

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24. The image below shows mRNA hybridized with the template DNA from which it was
transcribed. Label the following terms on the diagram: (5 points)

1.) DNA
2.) RNA
3.) Intron
4.) Poly A tail
5.) Location of the 5’ cap

introns

DNA

RNA Poly A
5’ cap tail

If the RNA and DNA strands were mixed up, half credit given for cap and polyA tail if shown on
the indicated RNA strand.

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25. You are conducting splicing experiments on gene X. The normal, wild-type (WT) RNA is
drawn below and shows characteristic splicing products and intermediates in the absence and
presence of okadaic acid, a splicing inhibitor. You have also generated the same RNA from a
cancer cell line reported to be defective in gene X splicing. The splicing intermediates for the
cancer derived RNA are shown below on the gel (mutant + okadaic acid). To the right of the gel
below, draw the splicing intermediates for the slowest migrating bands. Using the RNA diagram,
indicate the region of the RNA that is mutated in the cancer cells. (5 points)

The 3’ splice site in the intron is mutated.

Exon
2

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26. Answer the following questions about regulation of the lac operon in E. coli. (10 points)

The lac O operator sequence overlaps a portion of the lac operon promoter. Why is this
configuration important for the regulation of the lac operon?

Repressor has to block RNA polymerase binding to the promoter.

(Also accepted that this would block RNA polymerase from proceeding.)

How does an inducer work to turn on transcription of a bacterial operon that has been repressed,
such as the Lac operon?

Inducers bind to repressor inducing a conformational change in repressor, so it no longer binds

(it falls off) the region of DNA it is bound to.

When E. coli cells are growing in the presence of lactose with very little glucose, is the level of
cAMP high or low? Would the lac operon be transcribed?

cAMP levels would be high and the lac operon would be transcribed.

If the lac operon promoter sequence was mutated so that it could not bind RNA polymerase,
what would be the level of of transcription in the presence of very little glucose and high lactose?
Would cells be blue or white if they were grown in the presence of X-GAL?

It would not be transcribedà no expression. Cells would be white.

If the lacI gene was mutated so that the protein could no longer bind to lactose, what would be
the level of lac operon transcription in the presence of low glucose and high lactose? Would
cells be blue or white if they were grown in the presence of X-GAL?

Not expressed in either case since lacI is not expressed (is not produced) Cells would be white in
both cases.

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27. Pictured below is the DNA surrounding Gene Z. Expression of Gene Z is influenced by two
small molecules, pactose and yactose, and two proteins, Protein X and Protein Y. Protein X is a
repressor that only binds to DNA in the absence of pactose. Protein Y is an activator that only
binds to DNA in the presence of yactose.

In addition to the wildtype strain, you obtain a mutant that no longer produces Protein X. In the
table below, indicate when Gene Z will be expressed (using a “+” or “–“ to indicate your answer
is sufficient). (4 points)

Gene Z

Y-binding site X-binding site

Condition Wildtype X mutant

+pactose +yactose
+ +
+pactose -yactose
- -
-pactose +yactose
- +
-pactose -yactose
- -

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