Gen371 Sample Final Answer Key

Gen371 Sample Final Answer Key 1/26/11 4:19 PM
Home Sample Final Exam -- answers

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1. The following pedigree was constructed while mapping genes for coat color in dogs. A new locus, termed
Help hours Black , is thought to be responsible for black fur in these dogs. This locus is separate from previously described
loci and its exact map location is unknown. Answer the following questions using B and b for dominant and
Calendar recessive alleles, respectively, as appropriate.
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(a) I-1 was shown to be homozygous for the dominant black allele. Show the cross(es) and expected result(s)
that could have led to this conclusion. You are not limited to this pedigree.
Testcross I-1 (e.g., with I-2); all progeny should be black. (If I-1 were
heterozygous, that cross would yield black and non-black in 1:1 ratio.)
Note: You can't assume that just because some polymorphic markers are
homozygous, the Black gene is homozygous also -- we don't know beforehand
that the Black gene is contained within this region.
(b) Each individual in this pedigree was genotyped and phase was determined at 4 markers (E-H) on the same
chromosome as the Black locus. Each haplotype is shown vertically, but the origin of the haplotype (i.e.,
from the egg or sperm) is not shown. Both I-1 and I-3 were known to be homozygous for the dominant
black allele.
In the empty boxes below dog III-12, indicate which haplotype in that dog was maternally derived
(M) and which was paternally derived (P).
Draw a box around every haplotype or any part thereof that unambiguously was inherited from
either I-1 or I-3 by any of their descendants.
Based on this information, where could the Black locus be? (e.g., below Marker X, above Marker Y,
between X and Y, not on this chromosome. Note that there are no Markers X and Y, those are just
given as examples.) List all individuals supporting your conclusion.
Location of Black locus: Below E, above F Correction: above H
Individuals supporting your conclusion: III-4 and III-9 exclude E and above; III-6
excludes H and below.
Notes:
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This is not an association study, it is linkage analysis based on

recombination breakpoints. I-1 and I-3 are effectively unrelated
individuals, so the fact that they both have the same genotype at
markers F and G shouldn't be taken to mean anything. See the last
question on the cystic fibrosis homework in QS8.
(c) Based on sequence analysis of the region, two candidate genes for the Black locus were identified (genes 1
and 2). Northern blot analysis was done on mRNA from various tissues in dogs I-1 and I-2 , with Gene 1
DNA or Gene 2 DNA as probe. B = blood, S = skin, C = cheek.
Which gene is a better candidate for the Black locus? BRIEFLY explain how the northern blot
results fit with the observed fur phenotypes, including one explanation for the difference between
the B and b alleles.
Gene 2 is a better candidate--it shows a difference in skin mRNA

size between a black and a non-black dog. Possibly a splice
site difference (resulting in exons getting skipped during
splicing) or a deletion that has occurred in non-black dogs,
leading to a shorter mRNA in non-black dogs that presumably
doesn't code for functional protein.
Notes:
It isn't sufficient just to say that there is a difference in

Gene 2 mRNA; the tissue matters. Your answer has to give an
explanation for the size difference in mRNA and for the
difference in fur phenotype.
Suppose that the gene you rejected as a candidate is actually the Black gene. Suggest one way you
could then explain the northern blot results seen above for that gene.
Could be a point mutation in the coding sequence or some

mutation in an untranslated region of the mRNA so that the mRNAs
are the same size and abundance in black vs. non-black dogs, but
encoded protein sequence or amount is different, resulting in
the fur color difference.
2. A little-known species of yeast, Starbuckia u’villageana, has the remarkable property that it can grow on
caffeine as its sole carbon source. A researcher is testing two genes ( E and R ) for their involvement in caffeine
utilization. She knocks out each gene with Gen R , a dominant marker that confers resistance to the drug
Geneticin. Ultimately, she creates a diploid strain of genotype EeRr (where lower case = gene that has been
replaced by Gen R ). The diploid is able to grow on caffeine. She sporulates the diploid and collects tetrads of
spores, all of which are able to grow on complete plates.
(a) It is not known if these genes show complementation or have redundant functions. Depending on which is
true, tetrads could show different growth patterns on appropriate plates.
Suppose you look at the phenotypes of the haploid spores in ONE tetrad ( Tetrad 1 ) and the results
don’t let you distinguish between the two possibilities (complementation vs. redundant functions).
Fill out the diagram below, giving your predicted spore genotypes of such a tetrad and shading the
colonies to indicate predicted growth of those spores on the various plates. (Colonies on complete
plates have already been shaded.)
Similarly for Tetrad 2 , predict a tetrad outcome where it would be possible to tell if E and R show
complementation or are redundant. (Note: at this stage you are just making predictions of possible
outcomes. The actual results are given in part ‘b’ below.)
Notes
Reminder: if the genes complement, it means that functioning E and

functioning R are both needed for growth on caffeine. If they are
redundant, it means that it only takes one of the two genes to be
functional for the spore to grow on caffeine.
This problem can be approached a number of ways, but a simple one is just
to think of the possible spore genotypes. There are only four possible
spore genotypes from this diploid: ER, er, Er, and eR. A spore of
genotype ER will show growth on caffeine plates whether the two genes
complement or are redundant. A spore of genotype er will not grow on
caffeine in either scenario. Spores that are Er or eR will not grow on
caffeine if the two genes complement, but will grow if they are
redundant. So, a tetrad consisting only of ER and er spores will not let
you distinguish between complementation and redundancy; the two other
types of tetrads will.
(b) Eventually, 2000 spores from this diploid are examined; 1110 of them are able to utilize caffeine as their
sole carbon source. Assuming that E and R show redundant function, draw a genetic map for the two
genes that best fits the data. If you think there isn’t linkage, depict that in your answer. Show your logic,
including the genotype of the diploid (with the correct phase if appropriate).
For independent assortment of redundant genes, we expect 75% of spores

(genotypes ER, Er, and eR; =1500/2000) to show growth on caffeine.
Only 1110 show growth, so the deficit indicates (a) that the genes are
not assorting independently, and (b) that one of the parental types must
be er (the genotype that doesn't allow growth on caffeine).
Non-growth = one parental type (er) = 890, so total parental types =

1780.
Total recombinant type = 220; map distance = 220/2000 x 100 = 11 cM.
Linkage map (with phase as present in the diploid):
Notes
This problem should seem familiar -- it's basically the same problem that
you saw as a previous practice(the flagella genes). The species name has
been changed and the genes are redundant instead of complementing, but
the logic needed to solve it is the same.
3. Some zebrafish researchers decided to try positional cloning of the copper gene based on a dominant mutation
(cop) that makes bright copper-red fish (wild type fish are yellowish brown).
(a) The gel diagram (above, left) shows the copper genotypes and genotyping results (PCR products) for
polymorphic site z928 on the fish labeled "Fish1" through "Fish3". Two of these three fish were mated to
each other to produce Fish4.
Who were the parents of Fish4? __Fish1 and Fish2_______

Fill in the copper genotype of Fish4 in the box above that gel lane.
(b) Fish3 was mated to Fish4 and 2000 progeny were collected. In the blank gel images and spaces above, fill
in the expected number, phenotypes ("+" for wild type, "cop" for mutant phenotype), and z928 genotypes
(on the gel) if…
the copper gene does not show linkage to z928

copper and z928 are linked at a distance of 10 cM
Explanation:
Considering the Fish3 x Fish4 mating... Fish3 contributes only gametes of genotype + A so all the progeny
(the gels on the right) must have the A polymorphic allele. Fish4 is heterozygous cop/+ B/C . The parental
type gametes made by this fish are cop B and + C (because those are the gametes this fish received from
its parents). So the parental type gametes made by this fish will be cop B and + C; the non-parental
gametes will be cop C and + B . For independent assortment, these four gamete genotypes will be equally
probable (500 of each kind = 2000 total). If the cop locus is linked to the z928 polymorphic site at a
distance of 10 cM, the parental types will add up to 90% of 2000 = 1800, or 900 of each type. The non-
parentals will be 100 of each.
(c) After the researchers found a polymorphic site that was linked to the copper gene, they used PCR against a
genomic DNA library to identify a clone that contained that polymorphic site. They then tested this clone
for the presence of the copper gene by injecting it into fish embryos. (Injecting embryoes with DNA is way
of doing transformation in zebrafish.)
• What genotype do you think they used as the source of the DNA for the genomic DNA library?
(homozygous) copper fish [has the dominant allele]
• What is the genotype of the embryos they must have injected with the clone?
wild type [has recessive allele only]
• What specific result do you think they were hoping to get from the injections?
injected embryos should show copper phenotype
[There is a deeper question hidden here... what were the investigators trying to do? What was the point of
the PCR experiment, and what was their thinking there?
And in case you were wondering, this was a real experiment done to identify a real gene and published in
December 2005; I have just changed the names of the gene and the markers.]
4. Drosophila emeraldia is a newly discovered fruit fly species. Wild type flies of this species have brilliant green
eyes. Researchers have isolated two mutant lines, Eye1 and Eye2. Each line has a mutation at only one locus
and is homozygous for the mutation. Eye1 flies have unpigmented (white) eyes, whereas Eye2 flies have pale
yellowish-green eyes.
(a) A cross between Eye1 and Eye2 flies gives wild type progeny ("Progeny A"). Are these mutations
dominant or recessive? What else can you conclude so far with respect to eye color in these flies?
Eye1 mutation: dominant or recessive ? Eye2 mutation: dominant or recessive ?
What else you conclude:
Eye1 and Eye2 flies are mutated in two separate genes, which show
complementation: wild type alleles of both genes are needed for wild type
eye color.
(b) A cross between Progeny A females and fully recessive males (Males "B") gives wild type, pale
yellowish-green, and white-eyed flies in 1:1:2 proportions, respectively. Males and females of each
phenotype are found in equal proportions. Explain these phenotypes and proportions, giving genotypes
and stating your conclusions about linkage/non-linkage. Use upper case (E1, etc.) for dominant and lower
case (e1, etc.) for recessive alleles.
The two genes are autosomal and assort independently; e1 (unpigmented) is

epistatic to e2.
The cross is: E1/e1 E2/e2 x e1/e1 e2/e2 giving equal proportions of
E1/e1 E2/e2 (wild type)

E1/e1 e2/e2 (pale yellow-green eyes)
e1/e1 E2/e2 (unpigmented eyes)
e1/e1 e2/e2 (unpigmented eyes)
--> a 1:1:2 phenotype ratio.
(c) The researchers isolated mRNA from eyes of various flies, made fluorescent cDNA, and hybridized the
labeled cDNA from each fly line to microarrays containing sequences representing all the predicted fruit fly
genes (one microarray per fly strain). Their results for five predicted genes (1-5) are shown. The circles
represent microarray spots, where each spot contains DNA corresponding to that gene. Shading indicates
hybridization of cDNA; darker shading indicates more hybridization.
Which gene (1-5) is mutated in which mutant?
Eye1 line is mutated in gene # ____2_____
Eye2 line is mutated in gene # ____5_____
Note: You are told that each line is mutated in only one gene, so
although Eye2 shows a change in mRNA levels of two genes, there is only
one mutation. Since Eye1 is mutated in gene 2, the decrease in gene 2
mRNAlevels seen in Eye2 must be a consequence of the mutation in gene 5.
Draw a pathway consistent with all the above data to explain eye pigmentation in this species:
Pathway:
Note:
The pathway diagram has to account for the wild type and mutant
phenotypes (including epistasis) as well as the regulation of Gene 2 by
Gene 5. Invoking a second step in the synthesis of pigment, mediated by
Gene 5 protein, is not necessary but is okay as long as the regulation of
Gene 2 by Gene 5 is also indicated in the diagram.
Alternative pathways are possible also, for example:
How does your pathway account for the variation in mRNA levels seen for Gene 2 in the different strains?
Gene 2 protein is needed to make pigment; Gene 5 protein either

upregulates transcription of gene 2 (or prevents degradation of gene 2
mRNA) so that in the absence of functional gene 5, there is just low
level of Gene 2 mRNA and therefore pigment production is decreased,
giving pale yellowish-green eyes.
In the alternative pathway shown, the pale yellow-green intermediate is

capable of repressing Gene 2 transcription. When Gene 5 is not
functioning, the concentration of the yellow-green intermediate builds up
and therefore transcription of Gene 2 is reduced compared to wild type.
5. A research group has been studying a form of colorectal cancer that shows familial aggregation. In two
pedigrees, they have found significant evidence of linkage of colorectal cancer susceptibility to markers at a
recombinant percentage of θ = 10 (see table).
LOD score at θ = 10
in...
Marker Chromosome
Family 1 Family 2
A 11 0.2 0.1
B 11 3.4 0.33
C 11 0.03 -1.3
D 11 -2.3 -1.4
E 18 -1.3 0.03
F 18 1.1 1.2
G 18 -3.2 4.1
H 18 -0.7 0.2
(a) In analyzing and following up on the data, they chose not to add up the LOD scores from the two families.
Why might they have made this decision?
Evidence of linkage to two separate chromosomes (and evidence against
linkage to marker G in Family 1) indicates that mutations in two separate

genes are being transmitted in the two families. So, it would be
inappropriate to combine the scores from the two families.
(b) To follow up on the observed linkage, the

researchers looked for additional
polymorphic markers as outlined on the
right.
Why do you think they were

looking for additional markers?
To map the disease gene

at higher resolution.
What was the point of step (i)?
To find potential
polymorphic sites they
could use for the
higher-resolution
mapping.
What do you think they would have

been using as template (step iii)?
Pooled genomic DNA from

several different
individuals (preferably,
individuals in the family being examined).
Which sites shown in the gel diagram would they have considered not useful? Why?
1, 4, 5, 7, 8, 9, and 11 are not polymorphic and therefore not

useful for linkage mapping.
Notes: It is important to keep the "big picture" in mind here. If we are

following up on linkage by looking for additional markers, the goal must
be to do more mapping (i.e., to get more detail). To do mapping using
markers, the markers have to be polymorphic (i.e., multiple alleles
existing in the population). We first have to find potential markers
(step i, search the database for places that have CA repeats and
therefore potentially could be polymorphic) and then determine which of
those potential markers actually are polymorphic -- which we can do by
PCR-testing candidate markers using DNA from a number of people as
template, to see which of those candiate markers are in fact polymorphic
(step iii).
(c) Having identified a candidate gene on chromosome 11, the researchers genotyped a number of SNPs in the
region surrounding their candidate gene in cancer vs. healthy cells from a number of patients in the family
showing inheritance of susceptibility to colon cancer. One such example is shown below—genotyping was
done by allele-specific oligonucleotide hybridization; the SNPs tested are spread out over thousands of
base pairs. Shading indicates hybridization.
Explain the results...
What specific event do you think occurred in the cells that led to this cancer?
Most likely a deletion (one endpoint between SNPs 22 and 23, the
other endpoint between SNPs 27 and 28) of the wild type copy of
the gene, leading to loss of heterozygosity.
Write down the haplotype that is associated with cancer susceptibility in this individual. To indicate
uncertainty about the allele at a particular locus, write the alleles one above the other. For example,
if a haplotype could have either C or A at a position, you would indicate that as .
(Note: The first position should be read as "C or G", the second
position is "A or T", etc. Just saying TCGTC was okay also.)
Based on these results, do you think this cancer susceptibility gene is behaving as an oncogene or
more like a tumor suppressor gene? Explain.
An apparent deletion, which most likely would result in loss of

function, leads to cancer; the results better fit a model where
the cancer susceptibility gene is a tumor suppressor.
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Gen371 Sample Final Answer Key 1/26/11 4:19 PM

Home Sample Final Exam -- answers

Send email to class

Location of Black locus: Below E, above F Correction: above H

This is not an association study, it is linkage analysis based on

Gene 2 is a better candidate--it shows a difference in skin mRNA

It isn't sufficient just to say that there is a difference in

Could be a point mutation in the coding sequence or some

Reminder: if the genes complement, it means that functioning E and

For independent assortment of redundant genes, we expect 75% of spores

Non-growth = one parental type (er) = 890, so total parental types =

Total recombinant type = 220; map distance = 220/2000 x 100 = 11 cM.

Linkage map (with phase as present in the diploid):

Who were the parents of Fish4? __Fish1 and Fish2_______

the copper gene does not show linkage to z928

(homozygous) copper fish [has the dominant allele]

wild type [has recessive allele only]

injected embryos should show copper phenotype

Eye1 mutation: dominant or recessive ? Eye2 mutation: dominant or recessive ?

What else you conclude:

The two genes are autosomal and assort independently; e1 (unpigmented) is

E1/e1 E2/e2 (wild type)

--> a 1:1:2 phenotype ratio.

Which gene (1-5) is mutated in which mutant?

Eye1 line is mutated in gene # ____2_____

Eye2 line is mutated in gene # ____5_____

Alternative pathways are possible also, for example:

Gene 2 protein is needed to make pigment; Gene 5 protein either

In the alternative pathway shown, the pale yellow-green intermediate is

Evidence of linkage to two separate chromosomes (and evidence against

linkage to marker G in Family 1) indicates that mutations in two separate

(b) To follow up on the observed linkage, the

Why do you think they were

To map the disease gene

What was the point of step (i)?

What do you think they would have

Pooled genomic DNA from

1, 4, 5, 7, 8, 9, and 11 are not polymorphic and therefore not

Notes: It is important to keep the "big picture" in mind here. If we are

Explain the results...

An apparent deletion, which most likely would result in loss of

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Who were the parents of Fish4? Fish1 and Fish2_____

Eye1 line is mutated in gene # 2_

Eye2 line is mutated in gene # 5_