Zoo/Bot 3333
Genetics
Quiz 4
11/19/07
For answers to the quiz, click here:
Questions 1-4 pertain to the following. A mouse null mutant for a particular enzyme has been
identified by the fact that heterozygotes for the mutation produce ½ the amount of enzyme as normal
mice. By comparing mutant to wild type proteins on electrophoretic gels, the enzyme protein is
identified, eventually leading to the biochemical isolation of the enzyme protein. A small portion of
the sequence of the purified enzyme protein is determined to be:
Met Gly Ser Ala Gln Tyr Met Phe Asp Glu Trp Ser Met Ile Leu
This information is used to synthesize 21 base ‘degenerate’ oligonucleotides that will be used to screen
a cDNA library by nucleic acid hybridization for the cDNA encoding this particular enzyme.
1. What region of the amino acid sequence above should be used to manufacture an appropriate
‘degenerate’ probe?
a) Met-Gly-Ser-Ala-Gln-Tyr-Met; b) Ser-Ala-Gln-Tyr-Met-Phe-Asp; c) Gln-Tyr-Met-Phe-Asp-GluTrp; d) Asp-Glu-Trp-Ser-Met-Ile-Leu; e) any of the above would be equally effective.
2. How many different oligonucleotides would be present in the above degenerate probe?
a) 1; b) 7; c) 12; d) 32; e) none of the above.
3. What was the source molecule used to synthesize the cDNA that is present in the library to be
screened?
a) genomic DNA; b) mRNA from a tissue showing high levels of enzyme activity; c) mRNA from a
mouse that is homozygous mutant for the enzyme; d) protein purified from an electrophoretic gel; e) a
plasmid isolated from E. coli.
4. True or false. A single stranded degenerate probe encoding the protein sequence indicated above
could be also be used for a Northern blot analysis.
8 kb
4.5 kb
3.5 kb
Questions 5-6 pertain to the above: A large family is segregating an autosomal dominant disease of
late onset (approximately 40 years of age). A genomic DNA sample from each family member is
digested with the restriction enzyme HindIII, run on an electrophoretic gel, and subjected to Southern
blot analysis. The probe used in this instance hybridizes to a DNA fragment linked to the disease
gene, which shows polymorphism for this restriction enzyme. The autoradiogram of this blot is shown
above, aligned with the family pedigree.
5. In the above example, which of the following are likely to be physically linked to the disease gene
allele in individual I-1?
a) The 3.5 kb fragment; b) the 4.5 kb fragment; c) both the 3.5 and 4.5 kb fragments; d) the 8 kb
fragment; e) all three of the above fragments.
6. Which of the following most accurately represents the map distance between the disease gene and
the marker (probe DNA) locus?
a) 3.7 map units; b) 7.8 map units; c) 11 map units; d) 15.4 map units; e) 22 map units.
Consider the gel at the right, derived from a sequencing reaction based on the Sanger chain termination
method.
A
G
C
T
7. The sequence of the TEMPLATE STRAND
would be:
a) 5'-ACGTGCACGTGC-3';
b) 5'-CGTGCACGTGCA-3';
c) 5'- TGCACGTGCACG-3';
d) 5'-GCACGTGCACGT-3';
e) 5'-GCTAGCGCTAGC-3'.
8. True or false. Of the four DNA polymerase reactions used for Sanger DNA sequencing, only one
contains dideoxyadenosine triphosphate.
9. The restriction enzyme, AvaI has the six base recognition sequence CPyCGPuG, where Py
stands for a pyrimidine, and Pu stands for a purine). On average, this enzyme might be
expected to cut a random DNA sample every ____________bp.
a) 64; b) 256; c) 1024; d) 1296; e) 4096.
10. The recognition sites for the restriction enzymes BamHI, XbaI and BglII are,
respectively:
BamHI:
XbaI:
BglII:
5’-GGATC C-3’
3’-C CTAGG-5’
5’-TCTAG A-3’
3’-A GATCT-5’
5’-AGATC T-3’
3’-T CTAGA-5’
where the arrows represent the cut locations for each strand of the palindromic site. Which
enzymes leave compatible ends that will facilitate ligation?
a) All of these enzymes leave ends that are compatible with ends generated by the others; b)
None of the enzymes produce compatible ends; c) Only BamHI and BglII fragments are
compatible; d) Only BamHI and XbaI fragments are compatible; e) only BglII and XbaI
fragments are compatible.