Transcript Chapter 4

Griffiths • Wessler • Carroll • Doebley
Introduction to
Genetic Analysis
ELEVENTH EDITION
CHAPTER 4
Mapping Eukaryote Chromosomes
by Recombination
© 2015 W. H. Freeman and Company
CHAPTER OUTLINE
4.1
4.2
4.3
4.4
4.5
4.6
4.7
Diagnostics of linkage
Mapping by recombinant frequency
Mapping with molecular markers
Centromere mapping with linear tetrads
Using the chi-square test to infer linkage
Accounting for unseen multiple crossovers
Using recombination-based maps in conjunction
with physical maps
4.8 The molecular mechanism of crossing over
A recombination-based map of one of the chromosomes of Drosophila
Why map the genome ?

Gene position important to build complex
genomes

To determine the structure and function
of a gene

To determine the evolutionary
relationships and potential mechanism.
Two types of maps ?
 Recombination-based maps*
 Physical maps
The observation
1905 William Bateson and R.C. Punnett
Red petals, round pollen X
(rr,ss)
Purple petals, long pollen
(RR,SS)
F1 Purple petals, long pollen (Rr,Ss)
F1 selfed
(Rr,Ss) X (Rr,Ss)
Results
284
Purple, long
21
Purple, round
21
red, long
55
red, round
72
Purple, round
72
red, long
24
red, round
Expected F2
216
Purple, long
Symbols and terminology
AB
A/a
A/a; B/b
A/a . B/b
alleles on the same homolog, no punctuation
alleles on different homologs, slash
genes known to be on different
chromosomes, semicolon
genes of unknown linkage, use a dot or period
Cis
Trans
AB/ab or ++/ab
Ab/aB or +b/a+
Thomas Hunt Morgan & Drosophilia
Red eyes, normal
(pr+/pr+ . vg+/vg+)
X
Purple eyes, vestigal
(pr/pr . vg/vg)
F1 Red eyes, normal wings (pr+/pr . vg+/vg)
Instead of selfing the population, he did a test cross.
Test cross
Red eyes, normal
(pr+/pr . vg+/vg)
X
Purple eyes, vestigal
(pr/pr . vg/vg)
1339 Red eyes, normal wings (pr+ . vg+)
1195 Purple eyes, vestigal (pr . vg)
151 Red eyes, vestigal (pr+ . vg)
154 Purple eyes, normal wings (pr . vg+)
Test cross
1339 Red eyes, normal wings (pr+ . vg+)
1195 Purple eyes, vestigal (pr . vg)
151 Red eyes, vestigal (pr+ . vg)
154 Purple eyes, normal wings (pr . vg+)
pr+
305/2839 = 10.7 percent
pr
vg+
vg
cis or trans ?
Initial cross
Red eyes, vestigal
(pr+/pr+ . vg/vg)
X
Purple eyes, normal
(pr/pr . vg+/vg+)
F1 Red eyes, normal wings (pr+/pr . vg+/vg)
Test cross with pr/pr . vg/vg
157 Red eyes, normal wings (pr+ . vg+)
146 Purple eyes, vestigal (pr . vg)
965 Red eyes, vestigal (pr+ . vg)
1067 Purple eyes, normal wings (pr . vg+)
pr+
304/2335 = 12.9 percent
pr
vg
vg+
Linked alleles tend to be inherited together
Crossing over produces new allelic combinations
Chiasmata are the sites of crossing over. Occurs in Prophase I of meiosis.
Occurs in Prophase I (tetrad stage)
Crossing-over of the
chromosomes.
A chiasma is formed.
Genetic recombination.
Microscopic evidence for chromosome
breakage and gene recombination
Harriet Creighton and Barbara McClintock, 1931
-studied two genes in corn (on chromosome 9) for seed color
(C and c) and endosperm composition (Wx and wx)
Wx
C
wx
c
Wx
wx
c
C
Page 133 Note the knob on the C end and the longer piece of chromosome on the Wx end.
Recombinants are produced by crossovers
For linked genes, recombinant frequencies are less than 50 percent in a testcross
Mapping by Recombinant Frequency
Morgan set his student Alfred Sturtevant to the project.
“In the latter part of 1911, in conversation with Morgan,
I suddenly realized that the variations in strength of
linkage, already attributed by Morgan to differences in
the spatial separation of genes, offered the possibility of
determining sequence in the linear dimension of a
chromosome. I went home and spent most of the night
(to neglect of my undergraduate homework) in
producing the first chromosome map.” Sturtevant
Frequency of
crossing over
indicates the
distance between
two genes on the
chromosome.
A map of the 12 tomato chromosomes
Map distances are additive.
Question
You construct a genetic linkage map by following
allele combinations of three genes, X, Y, and Z.
You determine that X and Y are 3 cM apart, and X
and Z are 3 cM apart, and that Y and Z are 6 cM
apart. These cM numbers are most likely based
on:
a. DNA sequencing of the region in question
b. Recombination
b.
Recombination frequencies
frequencies
c. Measuring the distance in a scanning EM
micrograph
d. Independent assortment
Question
•
Referring to the cM numbers in the last
question, what is the relative gene order of
these three genes?
a.
b.
c.
d.
Z-X-Y
a.Y-X-Z
Z-X-Y
b. Y-X-Z
X-Y-Z
a and b
Longer regions have more crossovers and thus higher recombinant frequencies
Double recombinants arising from two crossovers
Different gene orders give different double recombinants
Phenotypic ratios in progeny reveal the type of cross
Summary
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•
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Gene linkage
Crossing over
Recombinant mapping
Be sure to read p. 96-98 and p. 150-151 in
order to do Chi-Square (c2) tests for some
homework problems
Review Problems
1. A plant of genotype
is test crossed.
A
B
a
b
If the two loci are 14 m.u. apart, what proportion
of progeny will be AB/ab ?
43% AB, 43% ab, 7% Ab, 7% aB
Review Problems
2. A plant of genotype A/a . B/b is test crossed.
The progeny are
74 A/a . B/b
76 a/a . b/b
678 A/a . b/b
672 a/a . B/b
Explain.
A and B are linked in trans and are 10 m.u. apart.
Review Problems
3. You have analyzed the progeny of a test cross to a tetrahybrid.
The results indicate that
10% of the progeny are recombinant for A and B
14% for B and C
24% for A and C
4% for B and D
10% for C and D
14% for A and D
Provide a linear map for the chromosome.
Review Problems
3. You have analyzed the progeny of a test cross to a tetrahybrid.
The results indicate that
10% of the progeny are recombinant for A and B
14% for B and C
24% for A and C
4% for B and D
10% for C and D
14% for A and D
Provide a linear map for the chromosome.
|----------|----|----------|
A 10 B 4 D 10 C
Mapping with Molecular Markers
Chapter 4, continued.
•
•
•
•
SNPs-single nucleotide polymorphisms
SSLPs-simple sequence length polymorphisms
SSLPs also called VNTRs-variable number tandem repeats
Molecular markers can be used instead of phenotype to map
genes
What is a molecular marker
 SNP = single nucleotide polymorphisms
AAGGCTCAT
TTCCGAGTA
AAGACTCAT
TTCTGAGTA
• Silent SNPs
• SNP that cause phenotype
• SNP in polygenes
• SNP in intergenic regions
• RFLPs (restriction fragment length
polymorphisms)
RFLPs

SNPs that introduce a restriction enzyme site.
GGATTC
CCTAAG
EcoR1 site
GAATTC
CTTAAG
digest with EcoR1
RFLP analysis
Fig 4-15a
RFLP analysis
Fig 4-15b
RFLP analysis
Fig 4-15c
Simple sequence length polymorphisms
(SSLPs)
Also known as VNTRs (variable number tandem repeats)
Repeats of DNA sequence, with different numbers of
repeats occurring in different individuals; used for
DNA fingerprinting.
Minisatellites (DNA fingerprints)
– Repeating units of 15-100 nucleotides
Microsatellites (DNA fingerprints)
– Repeating units of 2-15 nucleotides
[e.g., ACACACACACACAC]
Minisatellites
Fig. 4-18
Microsatellites
Amplified by polymerase chain reaction.
primer 1
CACACACACACACA
CACACACACA
GTGTGTGTGTGTGT
GTGTGTGTGT
primer 2
St. M
M’
A microsatellite locus can show linkage to a disease gene
Alignment of physical and recombination maps
Summary
• Mapping using molecular markers
– SNPs, RFLP mapping
– SSLPs/VNTRs
• Minisatellites
• Microsatellites