PROBLEM SET

Chapter 6 -- Linkage and Mapping in Eukaryotes

P. 134, #1. A homozygous groucho fly (gro, bristles clumped above the eyes) is
crossed with a homozygous rough fly (ro, eye abnormality). The F1 females are
testcrossed, with the following offspring produced:
groucho 518
rough 471
groucho, rough 6
wildtype 5
-------
1,000
a) What is the linkage arrangement of these loci?
To determine the linkage arrangement of these two loci we need to calculate
the map distance between them. The two classes in highest frequency are the
non-recombinant classes and the two classes in lowest frequency are the
recombinant classes. Thus groucho, rough and wildtype are recombinants. To
calculate the map distance, we divide the total number of recombinants by the
total number of flies and then multiply by 100.
11
map distance = ------ x 100 = 1.1 m.u. or 1.1 cM
1000
b) What offspring would result if the F1 dihybrids were crossed among
themselves instead of being testcrossed?
This is a little complicated because we need to set-up a Punnett square but the
four gametic genotypes will not have the same probability of occurrence of
0.25 each, as would expect if we had independent assortment. The first thing
then is to estimate the probability of occurrence for each gametic genotype. To
do this we must remember that a map distance of 1.1 m.u. means that a
recombinant gamete will occur with a probability of 0.011. Thus the two
recombinant gametes (gro,ro and +,+) will occur with a combined frequency
of 0.011, which when divided evenly between them means that the frequency
of occurrence for gro,rois 0.0055 and that the frequency for +,+ is also 0.0055.
The combined frequency of occurrence for the non-recombinant gametes
(gro,+ and +,ro) is 0.989 (1 - 0.011), which when evenly divided between
them means that the frequency of occurrence for gro,+ is 0.4945 and that the
frequency of occurrence for +,ro is 0.4945. The other thing to remember is that
in fruit flies, males do not crossover, thus one of the dihybrids (the male) will
not produce any recombinant gametes, and our Punnett square is reduced to a
Punnett rectangle.

By summing the frequency of occurrence for each cell in the Punnett Square,
we can determine the expected frequency for each phenotype in the F2.
groucho
gro gro, + + 0.24755
gro gro, ro + 0.00275
------------------
total 0.2500

rough
+ +, ro ro 0.24755
+ gro, ro ro 0.00275
--------------------
total 0.2500

wildtype
gro +, ro + 0.24725
gro +, ro + 0.24725
gro +, + + 0.00275
+ +, ro + 0.00275
--------------------
total 0.5000
Based upon the expected frequency of occurrence of the flies and assuming
1000 offspring, we would expect to see in the offspring of a dihybrid cross
wildtype 500
rough 250
groucho 250

P. 134, #2. A female fruit fly with abnormal eyes (abe) of a brown color
(bis, bistre) is crossed with a wildtype male. Her sons have abnormal,
brown eyes; her daughters are of the wildtype. When these flies F1 are
crossed among themselves, the following offspring are produced:
sons daughters
abnormal, brown 219 197
abnormal 43 45
brown 37 35
wildtype 201 223
What is the linkage arrangement of these loci?
In this case the two loci are on the X-chromosome. We know this because we
see a difference in the expression of the traits between the sexes in the F1
generation. However, this does not affect the working of the problem because
the ratio of males to females for each phenotypic class is approximately 1:1.
We can simply pool the sexes for each phenotype. Thus
abnormal brown 416
abnormal 88
brown 72
wildtype 424
That two classes in highest frequency are the non-recombinants and the two
classes in lowest frequency are the recombinants. To calculate the map distance
between the two loci, we use the formula above such that
160
map distance = ------ x 100 = 16 cM
1000
P. 134, #3. In Drosophila, the loci inflated (if, small, inflated wings) and
warty (wa, abnormal eyes) are about 10 map units apart on the X
chromosome. Construct a data set that would allow you to determine this
linkage arrangement. What differences would be involved if the loci were
located on an autosome.
To construct the data set, we need to setup the crosses and then calculate the
expected occurrence of each phenotype in the F2 generation. We will cross an
inflated, warty female with a wildtype male. This will produce in the F1 a
heterozygous wildtype female and a hemizygous inflated, warty male. These
two individual will then be mated to produce the F2 generation. As the male is
hemizygous for recessive traits, this is essentially a testcross as the gamete
from the F1 female will determine the phenotype of the F2 offspring. Also,
there will be no difference in the frequency of expression of the traits between
males and females in the F2 generation. For a map distance of 10, in the F2
generation 10% of the offspring must be recombinants. Thus from 1000
offspring, we expect 100 to be either inflated or warty as these are the
recombinant conditions given that the P generation was either inflated and
warty or wildtype. Of the 100 recombinant offspring, we will evenly divide
them among inflated (50) and warty (50). The nonrecombinant offspring will
be divided evenly among warty and inflated (450) and wildtype (450). As these
traits are X-linked, we could also evenly divide each of the phenotypic classes
evenly between males and females. This would give us a data set that is shown
below.
sons daughters total
inflated, warty 225 225 450
inflated 25 25 50
warty 25 25 50
wildtype 225 225 450
If these traits were on an autosome, the difference would be that to get the F2
generation, the female would have to be backcrossed to a male that was
homozygous inflated and warty and could not be crossed to her brother in the
F1 generation. In the F2 generation, the data set would look like that shown
under the total column above.

P. 134, #4. A geneticist crossed female fruit flies that were heterozygous at
three electrophoretic loci, each with fast and slow alleles, with males
homozygous for the slow alleles. The three loci were got-1 (glutamate
oxaloacetate transaminase-1) amy (alpha-amylase), and sdh (succinate
dehydrogenase). The first 1,000 offspring isolated had the following
genotypes:
class 1) got-s got-s,amy-s amy-s,sdh-s sdh-s 441
class 2) got-f got-s,amy-f amy-s,shd-f sdh-s 421
class 3) got-f got-s,amy-s amy-s,sdh-s sdh-s 11
class 4) got-s got-s,amy-f amy-s,sdh-f sdh-s 14
class 5) got-f got-s,amy-f amy-s,sdh-s sdh-s 58
class 6) got-s got-s,amy-s amy-s,sdh-f sdh-s 53
class 7) got-f got-s,amy-s amy-s,sdh-f sdh-s 1
class 8) got-s got-s,amy-f amy-s,sdh-s sdh-s 1
What are the linkage arrangements of these loci, including map units? If
the three loci are linked, what is the coefficient of coincidence?
In this case only two classes are in highest frequency, which indicates that all
three loci are linked. The next step is to determine the gene order. To do this we
must compare the parental classes (highest frequency) to the double crossover
classes (lowest frequency). As the male parent is homozygous for the slow
alleles, we can easily determine the gamete donated to the offspring by the
heterozygous female.
For the parental classes these are:
got-s,amy-s,sdh-s and got-f,amy-f,sdh-f
For the double crossover classes these are:
got-f,amy-s,sdh-f and got-s,amy-f,sdh-s
A comparison of the gametes from the parental classes with the double
crossover classes shows that the locus amy is in the middle.
The next step is to calculate the map distance among the loci. To do this we
need a table as given below
number of recombinants between
got-amy amy-sdh got-sdh
---------------------------------------
11 58 11
14 53 14
1 1 58
1 1 53
----------------------------------------
totals 27 113 136
To calculate the distance between each locus, we use the same formula as
before: the number of recombinants between the loci divided by the total
number of flies and then times 100.
27
distance (got-amy) = ------ x 100 = 2.7 m.u.
1000
113
distance (amy-sdh) = ------ x 100 = 11.3 m.u.
1000
distance (got-sdh) = 2.7 + 11.3 = 14.0
To calculate the coefficient of coincidence we need to calculate the expected
number of crossovers and then divide the number into the observed number of
double crossovers. The expected frequency of double crossovers is given by
multiplying the probability of a crossover between got-amy (0.027) by the
probability of a crossover between amy-sdh (0.113). Thus the probability of a
double crossover is 0.027 x 0.113 = 0.0031. Then multiplying this number by
the total number of offspring (0.0031 x 1000) = 3.1, which is the expected
number of offspring. The coefficient of coincidence is given by dividing the
observed number of double crossovers by the expected number of double
crossover (2/3.1) = 0.65.

P. 134, #5. The following three recessive markers are known in lab mice: h,
hotfoot; o, obese; and wa, waved. A trihybrid of unknown origin is
testcrossed, producing the following offspring:
hotfoot, obese, waved 357
hotfoot, obese 74
waved 66
obese 79
wildtype 343
hotfoot, waved 61
obese, waved 11
hotfoot 9
a) If the genes are linked, determine the relative order and the map
distance between them.
b) What was the cis-trans allele arrangement in the trihybrid parent?
c) Is there any crossover interference? If yes, how much?
In this case only two classes are in highest frequency, which indicates that all
three loci are linked. The next step is to determine the gene order. To do this we
must compare the parental classes (highest frequency) to the double crossover
classes (lowest frequency). As one parent is homozygous for the recessive
alleles, we can easily determine the gamete donated to the offspring by the
trihybrid parent.
For the parental classes these are:
h o wa and h+ o+ wa+
For the double crossover classes these are:
h+ o wa and h o+ wa+
A comparison of the gametes from the parental classes with the double
crossover classes shows that the locus hotfoot (h) is in the middle. In addition,
an examination of the nonrecombinant gametes shows that all widltype alleles
are on the same chromosome in the trihybrid and all the recessive alleles are on
the other chromosome. This is a cis arrangement of the alleles in the trihybrid.
The next step is to calculate the map distance among the loci. To do this we
need a table as given below
number of recombinants between
obese-hotfoot hotfoot-waved obese-waved
-----------------------------------------------
79 74 79
61 66 61
11 11 74
9 9 66
-----------------------------------------------
totals 160 160 160
To calculate the distance between each locus, we use the same formula as
before: the number of recombinants between the loci divided by the total
number of flies and then times 100.
160
distance (obese-hotfoot) = ------ x 100 = 16.0 m.u.
1000
160
distance (hotfoot-waved) = ------ x 100 = 16.0 m.u.
1000
distance (obese-waved) = 16.0 + 16.0 = 32.0
To calculate the coefficient of coincidence we need to calculate the expected
number of crossovers and then divide the number into the observed number of
double crossovers. The expected frequency of double crossovers is given by
multiplying the probability of a crossover between obese and hotfoot (0.160) by
the probability of a crossover between hotfoot and waved (0.160). Thus the
probability of a double crossover is 0.160 x 0.160 = 0.0256. Then multiplying
this number by the total number of offspring (0.0256 x 1000) = 25.6, which is
the expected number of offspring. The coefficient of coincidence is given by
dividing the observed number of double crossovers by the expected number of
double crossover (20/25.6) = 0.78. The percent of interference is given by (1 -
cc) x 100. percent interference = (1 - 0.78) x 100 = 22%.

P. 136, #23. In people, the ABO system (A, B, O alleles) is linked to the
aldolase-B locus (al,al+alleles), a gene that functions in the liver.
Deficiency, which is recessive, results in fructose intolerance. A man with
blood type AB had a fructose intolerant, type B father and a normal type
AB mother. He and a woman with blood type O and fructose intolerance
had ten children, of which five were type A and normal, three were
fructose intolerant and type B, and two were type A and intolerant to
fructose. Draw a pedigree of this family and determine the map distance
involved. (Calculate a lod score to determine the most likely recombination
frequency between the loci.
The first step is to create the pedigree for this cross and label the chromosomes
with as much inofrmation as is possible. The man with type AB blood had a
father who was homozygous for fructose intolerance, thus the B allele that the
man has must have come from his father and is linked to a fructose intolerant
allele. His mother had AB blood and was fructose tolerant and thus must have
at least one fructose tolerant allele al+. In this case the A allele from the mother
is linked to an al+ allele on the same chromosome. This makes the man with
AB blood heterozygous for both loci. He marries a woman who had type O
blood and is fructore intolerant, thus she is homozygous recessive at both loci.
The cross (heterozygous dominant x homozygous recessive) is a testcross.

Of the ten children 5 inherit the chromsome with the A allele without any
recombination and 3 inherit the chromosome with the B allele without any
recombination. Two of the children inherited recombinant chromoses. To
calculate the map distance between the two loci we use the formula below such
that
# recombinants
map distance = ---------------- x 100
total #
2
map distance = ---- x 100 = 20 cM
10
To calculate the lod score we must first calculate two probabilities. One is the
probability of these offspring given a map distance of 20 cM and then the
probability of these offspring given independant assortment. To determine the
probability of these offspring given 20 cM distance between the loci we must
first calculate the probability of any given child. Given a map distance of 20
cM, then 80% of the children will be nonrecombinants and 20% will be
recombinats. Thus the probability of a type A and normal (nonrecombinant) is
0.40, and the probability of a type B and fructose intolerant (nonrecombinant)
is 0.40, and the probability of a type A and fructose intolerant (recombinant) is
0.10. To get the probability of this family we then multiply the probabilities on
each birth together.
prob(given 20 cM distance) = 0.40^5 * 0.40^3 * 0.10^2 = 0.0000065536

To get the probability of this family given independant assortment, we assign
the probabiliy of any given phenotype the value of 0.25. Given a dihybrid
testcross, each phenotype is equally likely. To get the probability of this family
we then multiply the probabilities on each birth together.
prob(given independant assortment) = 0.25^10 = 0.0000009536743164062

To calculate the lod score we then take the logarithm of the ratio of these two
probabilities.
lod = log ((prob given 20 cM distance)/prob given independant
assortment))
0.00000655360
lod = log ------------- = log 6.872 = 0.83
0.00000095368
Any lod score greater than zero indicates linkage, but it is considered strong
evidence for linkage when the lod score is greater than 3.

P. 136, #24. In the mouse, the autosomal alleles Trembling and Rex (short
hair) are dominant to normal and long hair, respectively. Heterozygous
Trembling, Rex females were crossed to normal, long-haired males and
yielded the following offspring:
Trembling, Rex 42
Trembling, long-haired 105
normal, Rex 109
normal, long-haired 44
a) Are the two genes linked? How do you know?
The cross is a dihybrid female testcrossed to a homozygous recessive male. If
the two loci were not linked, then they would assort independently and you
would see the four phenotypes in the offspring in a ratio of 1:1:1:1. In this case,
two classes are in much higher frequency than the other two classes. This could
be tested with a chi-square, which would reject the null hypothesis of
independent assortment. Thus, we can see that the two loci do not assort
independently and we must conclude that they are linked.
b) In the heterozygous females, were Trembling and Rex in cis or trans
position? Explain.
The two classes in highest frequency represent the parental or nonrecombinant
classes. This means that the parents of the dihybrid female were Trembling,
long-haired and normal, Rex. The chromosomes of these parents would be
homozygous for these traits.
Trembling, long-haired normal, Rex
Tr + + R
------------ x ----------
------------ ----------
Tr + + R
The F1 dihybrid female (Trembling, Rex) would have received one of each
chromosome from her parents, one chromosome from Trembling, long-haired
and 1 from normal, Rex. Thus her chromosomes look like this
Tr +
---------------
---------------
+ R
In this case, Trembling is on one chromosome and Rex is on the homologue.
As they are on opposite chromosomes, they are in the trans position.
c) Calculate the percent recombination between the two genes.
We know that Trembling, long-haired and normal, Rex are the parentals thus,
Trembling, Rex and normal, long-haired must be recombinants. To calculate
the map distance, we use the formula
number of recombinants
map distance = --------------------------- x 100
total number individuals

86
map distance = ----- x 100 = 28.7 m.u.
300
P. 136, #25. In corn, a trihybrid Tunicate (T), Glossy (G), Liguled (L) plant
was crossed with a nontunicate, nonglossy, liguleless one and produced the
following offspring.
Tunicate, liguleless, Glossy 58
Tunicate, liguleless, nonglossy 15
Tunicate, Liguled, Glossy 55
Tunicate, Liguled, nonglossy 13
nontunicate, Liguled, Glossy 16
nontunicate, Liguled, nonglossy 53
nontunicate, liguleless, Glossy 14
nontunicate, liguleless, nonglossy 59
a. Determine which genes are linked.
In this case, an examination of the numbers of each phenotype shows that four
classes are in essentially equal frequency. This indicates that not all of the loci
are linked. The problem now becomes one of identifying which loci are linked.
The next step is to look at the phenotypic ratios taking the data two loci at a
time. The first pair will be the tunicate and ligule loci.
Ignore the phenotype at the glossy locus.
Tunicate, liguless 58 + 15 = 73
Tunicate, Liguled 55 + 13 = 68
nontunicate, liguless 14 + 59 = 73
nontunicate, Liguled 16 + 53 = 69
These four phenotypes are in about equal frequency, which indicates
independent assortment between the tunicate and liguled loci, and thus these
two loci are not linked.
The second pair will be the tunicate and glossy loci. Ignore the phenotype at the
liguled locus.
Tunicate, Glossy 58 + 55 = 113
Tunicate, nonglossy 15 + 13 = 28
nontunicate, Glossy 16 + 14 = 30
nontunicate, nonglossy 53 + 59 = 112
These four phenotypes are not in equal frequency, in fact two classes are at
much higher frequency (these are nonrecombinants) and two classes are at
much lower frequency (these are recombinants). Thus, these data indicate that
these two loci are linked. As we know that the liguled locus is not linked to the
tunicate locus and that the tunicate locus is linked to the glossy locus, then we
can assume that the liguled locus is not linked to the glossy locus. We could
show this by the above method, if necessary.
b. Determine the genotypes of the heterozygote; be sure to indicate which
alleles are on which chromosome.
At the tunicate, glossy loci, the nonrecombinants were tunicate, glossy and
nontunicate, nonglossy (see table above). This indicates that the chromosomes
in the heterozygote had the following arrangement.
T G L
---------------- ----------
---------------- ----------
t g l
The T and G alleles are cis on the same chromosome, however, the liguled
locus is on a separate chromosome.
c. Calculate the map distance between the linked genes.
Tunicate, Glossy 58 + 55 = 113
Tunicate, nonglossy 15 + 13 = 28
nontunicate, Glossy 16 + 14 = 30
nontunicate, nonglossy 53 + 59 = 112
From this table, the number of recombinants is 58 and the total numbers of
individuals is 283.
58
map distance = ----- x 100 = 20.5 m.u.
283

P. 136, #26. In Drosophila, kidney bean eye (k), cardinal eye (cd), and
ebony body (e) are three recessive alleles. If homozygous kidney, cardinal
females are crossed to homozygous ebony males, the F1 offspring are all
wildtype. If heterozygous F1 females are mated with kidney, cardinal,
ebony males, the following two thousand progeny appear:
880 kidney, cardinal
887 ebony
64 kidney, ebony
67 cardinal
49 kidney
46 ebony, cardinal
3 kidney, ebony, cardinal
4 wildtype
a. Determine the chromosomal composition of the F1 females.
From the flies in the P generation ("kidney, cardinal" and "ebony"), we know
that in the F1 one chromosome will carry the alleles k,cd,+ while the other
chromosome will carry the alleles +,+,e.
b. Derive a map of the three genes.
In this case only two classes are in highest frequency, which indicates that all
three loci are linked. That next step is to determine the gene order. To do this
we must compare the parental classes (highest frequency) to the double
crossover classes (lowest frequency).
For the parental classes these are:
kidney, cardinal and ebony
For the double crossover classes these are:
kidney, ebony, cardinal and wildtype
A comparison of the phenotypes from the parental classes(for example,
"kidney, cardinal") with the double crossover classes (for example "kidney,
ebony, cardinal") shows that the locus ebony is in the middle, because it is the
one locus that does not match.
The next step is to calculate the map distance among the loci. To do this we
need a table as given below
number of recombinants between
kidney-ebony ebony-cardinal kidney-cardinal
-----------------------------------------------------
64 49 49
67 46 46
3 3 64
4 4 67
------------------------------------------------------
totals 138 102 136
To calculate the distance between each locus, we use the same formula as
before: the number of recombinants between the loci divided by the total
number of flies and then times 100.
138
distance (kidney-ebony) = ------ x 100 = 6.9 m.u.
2000
102
distance (ebony-cardinal) = ------ x 100 = 5.1 m.u.
2000
distance (kidney-cardinal) = 6.9 + 5.1 = 12.0
The map is this.
k e cd
---+------------+------------+-------
6.9 m.u. 5.1 m.u.


P. 137, #30. Hemophilia and color blindness are X-linked recessive traits.
A normal woman whose mother was color-blind and whose father was a
hemophiliac mates with a normal man whose father was color-blind. They
have the following children:
4 normal duaghters
1 normal son
2 color-blind sons
2 hemophiliac sons
1 color-blind, hemophiliac son
Estimate the distance between the two genes.

P. 137, #31. The results of an anlysis of five human-mouse hybrids for five
enzymes are given in the following table along with the human
chromosomal content of each clone (+ = enzyme of chromosome present; -
= absence). Dedue which chromosome carries which gene.
see your book for the large table (Table 1)


Table 2. Clone
---------------------
human enzyme A B C D E
-------------------------------------------------
gluthathione reductase + + - - -
malate dehydrogenase - + - - -
adenosine deaminase - + - + +
galactokinase - + + - -
hexosaminidase + - - + -
-------------------------------------------------
The problem is to determine which clones have which enzymes. For example,
gluthathione reductase is present in clones A and B, but absent from clones C,
D, and E. The next step is to determine from Table 1 which chromosome or
chromosomes are in clones A and B and also absent from clones C, D, and E.
An examination of Table 1 shows that only chromosome 8 is present in clones
A and B while being absent from clones C, D, and E. Thus gluthatione
reductase is on chromosome 8.
For malate dehydrogenase we need to determine which chromosome is present
in clone B and absent from clones A, C, D and E. The only chromosome that
fits this criterion (present in B and absent from all others) is chromosome 2.
Thus malate dehydrogenase is on chromosome 2.
By the same reasoning as outlined above; adenosine deaminase is on
chromosome 20, galactokinase is on chromosome 17, and hexosaminidase is on
chromosome 5.

P. 138, #32. You have selected three mouse-human hybrid clones and
analyzed them for the presence of human chromosomes. You then analyze
each clone for the presence or absence of particular human enzymes (+ =
presence of human chromosome or enzyme activity). Based on the
following results, indicate the probable chromosomal location for each
enzyme.
Table 1. Human Chromosomes
-------------------------------
Clone 3 7 9 11 15 18 20
---------------------------------------------
X - + - + + - +
Y + + - + - + -
Z - + + - - + +
---------------------------------------------


Table 2. Enzyme
--------------------
Clone A B C D E
----------------------------------
X + + - - +
Y + - + + +
Z - - + - +
----------------------------------
Enzyme A is on chromosome 11. We know this because enzyme A is found in
clones X and Y but not in clone Z (Table 2). An examination of Table 1 reveals
that chromosome 11 is the only chromosome in clones X and Y but not in clone
Z. As the presence and absence of enzyme A matches that of chromosome 11,
we conclude that enzyme A in on chromsomes 11.
Enzyme B is on chromosome 15
Enzyme C is on chromosome 18
Enzyme D is on chromosome 3
Enzyme E is on chromosome 7

Last updated on 22 August 1996.
Provide comments to Dwight Moore at mooredwi@esumail.emporia.edu.
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