Download What is the genetic basis of complex traits? One of the most

CH927 Quantitative Genomics
What is the genetic basis of complex traits?
One of the most enduring problems
in evolution and molecular biology
What is the genetic basis of complex traits?
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Lecture 1 (Mon 9:30-10:30): markers, maps
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Lecture 2 (Mon 11:00-12:00): QTL methods
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Wet-bench practical (Mon 13:15-16:15): data for QTL mapping
** bus leaves to go to Warwick HRI at 12pm **
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Lecture 3 (Tues 9:30-10:30): Alternative methods: association mapping
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Lecture 4 (Tues 10:45-11:45): eQTL mapping
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Workshop (Tues 14:00-17:00): eQTL analysis using R-QTL
Lecture objectives
By the end of this lecture you should be able to explain:
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Quantitative genetics: homozygotes, heterozygotes and inheritance
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The basis and features of quantitative vs. qualitative traits
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Why genetic markers are needed for QTL mapping
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How genetic maps are created
And know what you’ll be doing in this afternoon’s practical at Warwick HRI
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Many sequenced genomes
Huge cost!
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But still not easy to identify
the right genes
Some definitions in molecular genetics
Genetics: the study of inheritance and its variations
Gene: the segment of DNA involved in producing a protein
Locus: a region of the genome, commonly a gene
DNA
promoter
exon intron exon intron exon
DNA
Chromosome: A linear end-to-end arrangement of genes and other DNA,
sometimes with associated protein and RNA
Genome: the entire complement of genetic material in an organism
Homozygosis vs. Heterozygosis
Cross pollination
Self pollination
Plant A
♂
e.g. one pair
of chromosomes
Plant B
♀ ♀
♂
Meiosis
pair is split
re-association (F1)
Identical
chromosomes
Identical genes
homozygous
Different
chromosomes
Different genes
heterozygous
Also during meiosis: crossing over occurs
Diploid: pair of
chromosomes from crosspollination
Duplication of the
chromosomes
We can use this property to
localise the parts of
chromosomes involved in a
trait
Crossing-over
Separation of
chromosomes
at end of meiosis
Quantitative vs. Qualitative traits
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Qualitative traits follow ‘Mendelian’ inheritance
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Can predict the phenotype from the alleles carried
e.g. A locus for eye colour with 2 alleles, B and b
- four possible combinations:
BB
Bb
bB
bb
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Dominant allele: same phenotypic character when heterozygous or
homozygous (Brown eyes: Bb bB BB)
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Recessive allele: phenotypic effect is expressed in homozyous state but
masked in heterozygous (Blue eyes in bb only)
Qualitative trait characteristics
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For qualitative traits you can predict the phenotype from the alleles
being carried
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These traits are often encoded by single genes e.g. albinism
Quantitative trait characteristics
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‘Infinitesimal model’: genetic variation in a trait due to a large number
of loci, each of small effect
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Many genotypes can produce the same phenotype
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Quantitative traits often vary along a continuous gradient
e.g. height, skin colour
diseases such as cancer
disorders such as epilepsy
non-Mendelian inheritance
What is the genetic basis of complex traits?
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Complexity of these traits, esp. those involved in adaptation
probably arises from segregation of alleles at many interacting loci
= Quantitative Trait Loci (QTL)
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QTL effects are sensitive to the environment
Combination of molecular genetics and statistical techniques
are needed to identify where these QTLs are located
Quantitative trait characteristics
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No typical patterns of dominance and recessiveness
Locus contributions thought to be additive (assumed)
= polygenic, or quantitative inheritance
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This can be explained as Mendelian inheritance at many loci (n)
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The coefficients of the binomial expansion
of (a + b)2n will give the frequency of
distribution of all n allele combinations
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For a sufficiently high n, this binomial
distribution will begin to be normal
increasing
disease
threshold for disease to occur
Lecture objectives
By the end of this lecture you should be able to explain:
•
Quantitative genetics: homozygotes, heterozygotes and inheritance
•
The basis and features of quantitative vs. qualitative traits
•
Why genetic markers are needed for QTL mapping
•
How genetic maps are created
Objectives of QTL analysis
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The statistical study of the alleles that occur in a locus and the
phenotypes (traits) that they produce
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Methods developed in the 1980s, perform on inbred strains of any species
1. Score a population for (i) a trait, and (ii) distribution of genome markers
2. Associate occurence of a marker with the phenotype
What do you need for QTL analysis?
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(i) A large population of individuals that you can score for phenotypes
and genotypes: Recombinant Inbred Lines (RILs)
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(ii) A map of the genome to find out where you are
(find out which chromosome the QTL is on)
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(iii) Markers over the genome to pinpoint QTL location
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(iv) A way to compare identify which markers from
each parent have been inherited by the progeny
- features to distinguish sequence from different origins
A
x
B
Parents =
Homozygous
F1 =
Heterozygous
at all loci
crossing-over
(recombination)
F2 =
Heterozygous
at some loci
x
Many different individuals
are obtained & separately
selfed to develop RILs
x5
F7 RILs =
Homozygous
at all loci
& heterogeneous
(i) A large population of mapping Recombinant Inbred Lines
(ii) Markers to enable identification of which parental genome
each part of the chromosomes of the progeny have come from
parent A
parent B
parent A
parent B
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Visible phenotypes or molecular markers (DNA sequence differences)
(iii) A map of the genome: anchor the markers
Parent A
Chr 1
Parent A
Chr 2
Different chromosomes
Molecular markers = features of the DNA sequence
Markers differ between parents (natural variants)
Parent B
Chr 1
Parent A
Chr 1
Different species variants
single nucleotide polymorphisms
GAATTC
GATTTC
(iv) You can distinguish these sequence differences using
molecular techniques = molecular markers
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Restriction enzymes e.g. EcoRI cut DNA
only at a specific recognition sequence
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Compare restriction patterns:
Parent A
Parent B
........GAATTC.......GAATTC.......GAATTC.......
........GAATTC.......GATTTC.......GAATTC.......
........GAATTC.......GAATTC.......GAATTC.......
........GAATTC.......GATTTC.......GAATTC.......
First generation (F1)
Second generation (F2)
from selfing F1:
There are many types of molecular markers
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Restriction Fragment Length Polymorphisms (RFLPs)
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Simple Sequence Length Polymorphisms (SSLPs)
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Cleaved Amplified Polymorphic Sequences (CAPS)
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Microsatellites (repeated sequences of 1-6 bases)
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Essentially, all of these are methods with which to detect sequence
differences that have occured between two variants of a species
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They mostly differentiate single nucleotide polymorphisms (snps)
Lecture objectives
By the end of this lecture you should be able to explain:
•
Quantitative genetics: homozygotes, heterozygotes and inheritance
•
The basis and features of quantitative vs. qualitative traits
•
Why genetic markers are needed for QTL mapping
•
How genetic maps are created
Need to know the linkage order: making a genetic map
There are two types of maps:
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Physical map: lays out the sequence information
and annotates it: promoters, genes etc.
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a
A
B
b
Linkage map: order of genetic markers and relative
distances from each other
- plus how much meiotic recombination (crossing over)
there is between homologous chromosomes carrying
alternative alleles (genetic markers)
Genetic linkage is related to recombination frequency
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Are loci A and B linked (on same chromosome) or unlinked (different chromosomes)?
a
a
A
B
B
b
Rf = 0.5 (50%)
= no linkage
aB, Ab, ab, and AB
in equal proportions
a
B
A
b
a
B
A
b
More recombination
so Rf = high ( <0.5 )
= weak linkage
aB, Ab, ab, and AB
in similar proportions
Rf = recombination
frequency
A
b
Little recombination
so Rf = small
= tight linkage
Some recombination
so Rf = medium
= quantifiable linkage
More aB, Ab
than ab, AB
Only aB and Ab
a
B
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A
b
Map distances and genetic linkage
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Recombination frequency of 0.01 (1%) = a genetic map unit of 1 cM
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Recombination events occur randomly, once or twice per chromosome
A linkage map is made by characterising the recombination events that
have taken place in a cross between two parental genotypes
** Every individual cross will have an individual linkage map **
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To make a map you need to score many markers in many individuals
• Assumes that linkage is the only cause of non-independence between
markers and that segregation is Mendelian
a
B
Determining map order
A
b
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Likelihood ODds ratio: likelihood of the observed linkage
The higher the LOD score, the more closely linked the markers are
Data on the presence/absence of 100s of markers in (F7) progeny population
Then you can use statistics to work out the marker order
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Traditionally done by hand using e.g. the Chi-squared statistic to test for
goodness of fit for the observed segregation ratios between markers
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With even just 10 marker scores, this means looking at many combinations:
1 2 3 4 5 6...
1 3 2 4 5 6...
1 3 4 2 5 6... and so on...
= (10 x 9 x 7 x 6 x 5 x 4 x 3 x 2 x 1)/2 = 1,814,400 possible orders!!
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That’s a lot of Chi-squared tests!
So we use mapping software e.g. Mapmaker, JoinMap
Determining map order
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Recombination fraction = n recombinant gametes
total
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Haldane mapping function adjusts map distance to account
for double crossovers that go undetected
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Map distance ≈ (RAB + RAC - 2RABRBC) x 100 cM
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2RABRAC is negligible for <10cM
A
a
B
b
C
c
Kosambi mapping function also adjusts for crossover interference
i.e. a crossover reduces the probability of a second crossover nearby
Linkage groups are the basis of genetic maps
These should theoretically correspond to chromosomes, but if...
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Chromosomes very long
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Recombination frequency very high
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Mapping populations are not large enough
...one chromosome can statistically “break” into several linkage groups
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Also, centromeres and heterochromatin have supressed recombination
A genetic linkage map for broccoli
1
2
3
4
5
6
7
8
map units
cM
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Recombination frequency of 0.01 (1%) = a genetic map unit of 1 cM
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