Download Lecture 5: Linkage and gene mapping

Lecture 5: Linkage and
gene mapping
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Two point test cross - review
Discovery of Linkage
Cross-over
Meiotic recombination
Gene mapping by
two-point testcross
Thomas H. Morgan
Parental
gametes
Gametes
1. Two types of gametes:
parental and recombinant
dissimilar to
either parental
type gamete
Parental
gametes
Gametes
1. Two types of gametes:
parental and recombinant
2. Equal frequency due to
independent assortment
1/4
1/4
1/4
1/4
Two-point test cross
(always a homozygous
recessive!)
A testcross in Drosophila that involved
two autosomal genes:
eye color - pr, purple, pr+, red
wing length - vg, vestigial, vg+, normal
Wild type alleles are dominant
vestigial mutant
wild type
P:
Gametes:
pr / pr . vg / vg (double mutant)
X
pr+ / pr+ . vg+ / vg+ (wild type)
pr+ vg+
pr vg
pr pr+ vg vg+
F1
testcross of F1 ♀:
Gametes of F1 ♀
pr vg
pr+ vg+
pr+ vg
pr vg+
pr pr+ vg vg+
X
Gametes of ♂
pr vg
“
“
“
♂ pr pr vg vg (homozygous recessive)
Classes of F2
pr pr vg vg
pr+ pr vg+ vg
pr+ pr vg vg
pr pr vg+ vg
Observed results
1195
1339
151
154
Four classes in F2, but not in 1:1:1:1 ratio!
Morgan’s interpretation of the results:
• obviously, no independent assortment of pr+/pr
and vg+/vg pairs of alleles
• the purple and vestigial genes are ‘linked’ to
each other on the same chromosome, each gene
residing in its own locus (position)
• in one parent, the specific alleles pr and vg are
linked, and in the other parent the alleles pr+ and
vg+ are linked
• in heterozygous F1, these alleles reside in the
different homologous chromosomes
• the same alleles preferentially remain together
in gametes of F1 forming two most abundant
classes of gametes
• these gametes form most abundant F2 progeny
P:
Gametes:
Most
abundant
(parental type)
gametes of F1
pr / pr . vg / vg (double mutant)
X
pr+ vg+
pr vg
pr pr+ vg vg+
F1
testcross of F1 ♀:
Gametes of F1 ♀
pr vg
pr+ vg+
pr+ vg
pr vg+
pr+ / pr+ . vg+ / vg+ (wild type)
pr pr+ vg vg+
X
Gametes of ♂
pr vg
“
“
“
♂ pr pr vg vg (homozygous recessive)
Classes of F2
pr pr vg vg
pr+ pr vg+ vg
pr+ pr vg vg
pr pr vg+ vg
Observed results
1195
1339
151
154
If this is true, we can predict the two most abundant classes in a cross with the same pair of
alleles but in a different configuration:
pr+ / pr+ . vg / vg (vestigial mutant)
P:
pr / pr . vg+ / vg+ (purple eyes mutant)
x
pr+ vg
Gametes:
pr vg+
+
pr+ pr vg+ vg
F1:
Testcross of F1 ♀:
pr+ pr vg+ vg
x
♂ pr pr vg vg
Gametes of F1 ♀
pr vg
pr+ vg+
pr+ vg
pr vg+
Gametes of ♂
pr vg
“
“
“
Classes
of F2
pr pr vg vg
pr+ pr vg+ vg
pr+ pr vg vg
pr pr vg+ vg
Observed results
146
157
965
1067
¾ In each cross there are two major and two minor classes
¾ The major classes represent those that are formed by F1 female gametes with
the original (Parental) configuration of alleles:
in the cross above these are:
pr+ vg
and pr vg+
in the previous cross these are:
pr vg and pr+ vg+
But what is the origin of the minor classes?
Chromosomal cross-overs
occur during meiosis in Prophase I, visible in Metaphase I as
chiasma (plural - chiasmata)
Recombinant
chromosomes
(parental gametes)
(in F1)
(in gametes of F1)
Formation of recombinant gametes by a cross-over during
meiosis in F1 heterozygotes
pr+
pr+
pr
vg+
pr+
vg+
vg
pr
vg
Before
Meiosis I
Prophase I:
cross-over
pr+
vg+
pr+
vg
pr
vg+
pr
vg
vg+
Parental
pr+
vg
pr
vg+
pr
vg
Recombinant
Recombinant
Parental
After Meiosis I
After
Meiosis II
Recombination is a process which generates genotypes different from the
parental genotypes
However, many meioses, there will be no cross-over
between the two loci. Only parental type gametes will be
formed
pr+
pr+
vg+
Parental
vg+
pr+
vg+
pr+
vg+
pr+
pr
vg
pr
vg
pr
vg+
Parental
vg
Parental
Before
Meiosis I
Prophase I:
cross-over
pr
vg
pr
vg
Parental
After Meiosis I
After
Meiosis II
Overall, there will be more parental type gametes (hence, parental type
progeny) than recombinant gametes
Observed frequency of crossovers reflects a distance between
the loci
Gametes:
1. The locus A and the locus B are very
close. When a crossover happens, it will
most likely happen outside the distance
between the loci, and no recombinant
gametes for the alleles in question will be
produced
2. The locus A and the locus B are very far
apart. Any crossover that happens, will most
likely happen between the loci. A half of all
gametes will be recombinant, and a half will
be parental.
A B
a b
b
a
A
3. The locus A and the locus B are at some
intermediate distance. In some meiocytes, a
crossover would occur in between the loci
generating both recombinant and parental
gametes. But in other meiocytes, a crossover would occur elsewhere, generating only
parental gametes. Overall, less than a half of
the gametes will be recombinant.
B
A
AB
AB
ab
ab
Parental = 100%
Recombinant = 0%
AB
Ab
aB
ab
Recombinant = 50%
(Parental = 50%)
B
a
A
b
B
a
b
AB
AB
ab
ab
AB
Ab
aB
ab
0%< Recombinant <50%
(100%> Parental > 50%)
Note: recombination between two loci can only be observed if a crossover occurs in
between the two loci, not just somewhere along the chromosome
Recombination frequency is a function of map distance
The plot is linear
approx. to RF ~ 7%
RFmax = 50%
Meiotic recombination
Interchromosomal: Mendel’s
Independent Assortment
Recombination frequency is 50%
Intrachromosomal: Morgan’s cross-overs
between linked loci
Recombination frequency is <50%
Linkage mapping by
two-point testcross
Locus A
Locus B
X map units
1 map unit (centiMorgan) =
1 % recombination frequency
X%
Let’s assume it is 5 m.u.
that is RF (A, B) = 5%)
Locus A
Locus B
5 map units
1 map unit (centiMorgan) =
1 % recombination frequency
5%
Let’s put the third locus
(locus C) on the map.
How to determine the
distance
between A and C?
Let’s assume RF(A,C) = 3%
Locus A
C
c
C
c
C
c
Locus C
C
c
c
c
C
c
3 map units
c
c
c
c
3%
C
c
Putting the three loci together: determine RF (B,C)
…if the distance between loci
B and C is 8 m. u.
..then there is only one
map that conforms all
the distances
…the alternative map would
be correct if distance B and C
were 2 m. u.
Note: map distances are additive
Map distances determined in a single experiment
cannot exceed 50 m.u., but combined map
distances can be much larger
70 m.u.