Genetics Final, Young Fall ’09, 150 pts.
Name:______________________
Due midnight, 12/9/09 by email (subject line must include “321m”), -or- 4:30 in the Biology Office
1. (20 pts) You are a genetic counselor and are presented with a family in which a disease
is present as indicated below. You’ve isolated DNA from each family member shown and
have obtained PCR primers for the region surrounding a common SNP (114A>C, A
being the wt nucleotide). Cycle sequencing confirms that mutations in this family are
caused by this transition / transversion (underline the correct one, 2 pts) at nucleotide
114. You buy ASOs to detect sequence at 114.
1a. (2 pts) What is the mode of inheritance, based
on the pedigree?
1b. Target DNA from each individual was
obtained with PCR, and blotted to replicate blots
(blot 1 and blot 2). Blot 1 was hybridized with a labeled C ASO and blot 2 with a
labeled wt ASO. Fill in the circles on each blot that would show positive hybridization.
For circles for which there isn’t enough information, indicate the probability of it being
positive. (or indicate, by underlining, bold, etc. for email submission.)
blot 1
I1
II1
II2
II3
II4
II8
III8
IV1
IV2
blot 2
I1
II1
II2
II3
II4
II8
III8
IV1
IV2
2. (10 pts) Starting with no sequence information, use the following words only (and
comma), to succinctly describe how to identify a gene that functions in mouse
veromonasal tissue and clone it, including the promoter and all introns. You may use a
word more than once, or not at all.
mouse genomic transcriptase scan identify isolate make that hybridizes veromonasal create extract detect
cDNA tissue mRNA a probe library from laser clone target the reverse use to from , microarray
3. (12 pts) For the following questions, use the amino sequence, in tandem with the codon usage
table, on the Reference Page (it's the last page on your exam, and can be detached as you won't be
asked to hand it in). The 638 amino acid sequence represents the complete human growth factor
protein as deduced from a cDNA. Single letter amino acid designations refer to specific codons
(i.e. M = Met: AUG).
Note: Use the topmost codon if the code is degenerate at that position.
a. (2 pts) How many DNA base pairs code for the protein?
b. (6 pts) Design 12 base pair DNA primers that you can use to PCR amplify the entire gene.
5' - ___________________________ 3'
5' - ____________________________ 3'
c. (2 pts) How long (in base pairs) is the PCR product from 3b, given a complete cDAN acts
as the template?
d. (2 pts) You also have genomic DNA with the whole HGF gene. You use your PCR
primers on the genomic template and get a PCR product that is nearly twice as long as the
PCR product in 3c. I one word, why?
4. (16 pts) Consider Fig. 4. Detection of
viruses in isolated nasal lavage from Wang et
al., Microarray-based detection and genotyping
of viral pathogens, PNAS 99(24).
a. (4 pts) Is this an image of a (underline
correct answer)….
i. Microarray
ii. Microchip
iii. Neither
b. (6 pts) How was the target obtained in the RV16/HeLa Amplified portion of this
figure?
c. (6 pts) What was the probe used for the RV16 (+/-) section of the figure. How was it
chosen for inclusion in this experiment, and how was it made?
5. (18pts) Use the following sequenced DNA as a template for PCR:
5'-ATGCATGCGA GTAGTCGTGT AGTCGATGTA CTGCATGTTT ACGTCGTAGT GGTACGATGT
|||||||||| |||||||||| |||||||||| |||||||||| |||||||||| ||||||||||
3'-TACGTACGCT CATCAGCACA TCAGCTACAT GACGTACAAA TGCAGCATCA CCATGCTACA
61
91
AGTCGATGTA CTGCATGTTT AAGTCGTAGT AAGCTTATGT GTACGTGTGA TGCTGCTAGT
|||||||||| |||||||||| |||||||||| |||||||||| |||||||||| ||||||||||
TCAGCTACAT GACGTACAAA TTCAGCATCA TTCGAATACA CATGCACACT ACGACGATCA
131
TACTGACGTG TTACGTGCTA GTACGTGTGA TGCTGCTAGT GTACGTGCAT GTTACTTTGT-3'
|||||||||| |||||||||| |||||||||| |||||||||| |||||||||| ||||||||||
ATGACTGCAC AATGCACGAT CATGCACACT ACGACGATCA CATGCACGTA CAATGAAACA-5'
a. Circle a pair of 20 base sequences that would be used for PCR primers to
amplify a 100 bp DNA product containing the G/C base pairs at position 93.
b. (4 pts) Write the nucleotide sequence for each primer in 5' - 3' orientation,
Primer 1: 5' - __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ - 3'
Primer 2: 5' - __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ - 3'
c. (2 pts) Exactly how long will the predominant PCR product be after 41 rounds?
___________bp
d. (6 pts) Position 93 (G) is the third base in a codon (codon table on back page)
that codes for lysine, and has been found to be a highly polymorphic site that
when mutated may be lethal. Design 15 base pair ASOs that will hybridize to
DNA from individuals who carry a lethal mutation as heterozygotes, give the
mutant sequence that will be recognized (indicate 5’, 3’ orientation), and circle
the base that is diagnostic:
ASO (wt): 5' - __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ - 3'
ASO (mutant): 5' - __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ - 3'
Mutant Sequence:
____- __ __ __ __ __ __ __ __ __ __ __ __ __ __ __ - ____
e. (6 pts) The AAGCTT at positions 91-96 is a palindrome recognized by the
restriction enzyme HindIII. Draw the results of a Southern blot with DNA
subjected to HindII digestion, and hybridized with labeled wt DNA.
wt
100
80
60
bp
40
20
heterozygote
homo-zygote
mutant
6. (11 pts) Consider Fig. 3 Two-dimensional gel electrophoresis of proteins from the
cytosolic fraction of human from Intra- and Interspecific Variation in Primate Gene
Expression Patterns, Science(296). A. Human, B. Chimpanzee. Check all correct
answers…
6a. (4 pts) The spot labeled “5” in 3A and 3B (underline all correct answers),
…is expressed more highly in chimpanzees than in humans,
…is expressed more highly in humans than in chimpanzees,
…has different 1o sequence in the two species,
…has similar 1o sequence in the two species.
6b. (4 pts) The spot labeled “13” in 3A and 3B (underline all correct answers),
…is expressed more highly in chimpanzees than in humans,
…is expressed more highly in humans than in chimpanzees,
…has different 1o sequence in the two species,
…has similar 1o sequence in the two species.
6c. (3 pts) The “spots” are (underline all correct answers),
i. mRNA
ii. cDNA iii. Genomic DNA
iv. Proteins v. Probes vi. Targets
7a. (4 pts) Considering virus detection, what’s the difference between a primer used in a
multiplexed PCR reaction with a primer used in a degenerate PCR reaction?
7b. (4 pts) What’s the most common G6PD deficient mutant found in African
populations?
What are the amino acid changes?
8. (12 pts) Consider Fig. 1 Targeted deletion of a cluster of V1r genes from Deficient
pheromone responses in mice lacking a cluster of vomeronasal receptor genes, Nature
(419).
a. (4 pts) What are V1r genes?
b. (4 pts) Why wereV1ra,b genes selected for this
reverse genetic study?
c. (4 pts) On what basis were the V1r genes clustered for this diagram?
i. DNA array data ii. DNA sequence data
iii. DNA map data
iv. Behavior analysis
9. (12 pts) Consider Fig. 2. Distance trees representing the
relative extent of expression changes among three primate
species and three tissues as assayed by the DNA arrays
from Intra- and Interspecific Variation in Primate Gene
Expression Patterns, Science(296).
a. (4 pts) What was the target in this experiment?
___________________
b. (4 pts) What was the probe in this experiment?
i. cDNA,
iii. mRNA
ii. genomic DNA,
iv. all of these
Fig. 2
c. (4 pts) What is different between Fig. 1a and Fig. 2 in this paper?
i. probes are longer in Fig. 1
ii. probes are longer in Fig. 2
iii. targets are longer in Fig. 1
iv. targets are longer in Fig. 2
Fig. 1a.
10. (18 pts) Consider Fig. 1. Metabolic defect in hereditary fructose intolerance from
The genetic consequences of our sweet tooth, Nature Reviews Genetics (3).
a. (4 pts) Circle the main compound that
builds up in patients with HFI.
b. (4 pts) Underline the enzymatic step(s) in
this diagram that are genetically blocked in
persons with Hereditary Fructose Intolerance
(HFI).
c. (6 pts) Place stars beside compounds that
do not accumulate in patients with HFI,
compared with wt.
d. (6 pts) Fructokinase activity is eventually
inhibited by high levels of fructose-1-phosphate. After an HFI event, which of the
following sugars would be safe for the affected individual to consume (circle all possible
answers), if they want to get better?
i. Glucose
ii. Fructose
iii. Sucrose
iv. Lactose
11. (13 pts ) The following represents a genomic map of a mutant gene. The wild-type
version of the gene lacks the BamH1 site on the second exon. The probe homology
region is shown below the scale (100 bp per tick, all map points not falling on a tick are
at 50 bp). You have genomic DNA and full length cDNA, and cut them with BamH1.
Probe
wt
mut
ladder genomic genomic
wt
mut
cDNA
cDNA
You run the samples on a gel,
blot and probe. Complete this
blot by creating a ladder, and
indicating the locations of any
fragments that hybridize. If you
can’t draw for email, just tell
me the sizes.
Reference Page
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