Download - CH927 Quantitative Genomics What is the genetic basis of ...By the end of this lecture you should be able to explain: • Quantitative genetics: homozygotes, heterozygotes and inheritance

What is the genetic basis of complex traits?

One of the most enduring problemsin evolution and molecular biology

CH927 Quantitative Genomics


• Lecture 1 (Mon 9:30-10:30): markers, maps

• Lecture 2 (Mon 11:00-12:00): QTL methods

• Wet-bench practical (Mon 13:15-16:15): data for QTL mapping ** bus leaves to go to Warwick HRI at 12pm **

• Lecture 3 (Tues 9:30-10:30): Alternative methods: association mapping

• Lecture 4 (Tues 10:45-11:45): eQTL mapping

• Workshop (Tues 14:00-17:00): eQTL analysis using R-QTL

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

Lecture objectives

And know what you’ll be doing in this afternoon’s practical at Warwick HRI

• Many sequenced genomes

• Huge cost!

• But still not easy to identify the right genes

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

Some definitions in molecular genetics

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

Self pollination Cross pollination

Plant A Plant B

♂ ♀ ♀ ♂e.g. one pair

of chromosomes

re-association (F1)

pair is split Meiosis

Different chromosomesDifferent genesheterozygous

Identical chromosomesIdentical geneshomozygous

Diploid: pair of chromosomes from cross-

pollination

Duplication of the chromosomes

We can use this property to localise the parts of

chromosomes involved in a trait

Also during meiosis: crossing over occurs

Crossing-over

Separation of chromosomes

at end of meiosis

Quantitative vs. Qualitative traits

• Qualitative traits follow ‘Mendelian’ inheritance

• Can predict the phenotype from the alleles carried

• Recessive allele: phenotypic effect is expressed in homozyous state but masked in heterozygous (Blue eyes in bb only)

• Dominant allele: same phenotypic character when heterozygous or homozygous (Brown eyes: Bb bB BB)

e.g. A locus for eye colour with 2 alleles, B and b

- four possible combinations: BB Bb bB bb

Qualitative trait characteristics

• For qualitative traits you can predict the phenotype from the alleles being carried

• These traits are often encoded by single genes e.g. albinism

Quantitative trait characteristics

• ‘Infinitesimal model’: genetic variation in a trait due to a large number of loci, each of small effect

• Many genotypes can produce the same phenotype

• Quantitative traits often vary along a continuous gradient

e.g. height, skin colour diseases such as cancer disorders such as epilepsy

non-Mendelian inheritance


• Complexity of these traits, esp. those involved in adaptation probably arises from segregation of alleles at many interacting loci

= Quantitative Trait Loci (QTL)

• Combination of molecular genetics and statistical techniques are needed to identify where these QTLs are located

• QTL effects are sensitive to the environment

• No typical patterns of dominance and recessiveness• Locus contributions thought to be additive (assumed) = polygenic, or quantitative inheritance

Quantitative trait characteristics

threshold for disease to occur

increasingdisease

• The coefficients of the binomial expansion of (a + b)2n will give the frequency of distribution of all n allele combinations

• For a sufficiently high n, this binomial distribution will begin to be normal

• This can be explained as Mendelian inheritance at many loci (n)






Lecture objectives

Objectives of QTL analysis

• The statistical study of the alleles that occur in a locus and the phenotypes (traits) that they produce

• 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

• (iii) Markers over the genome to pinpoint QTL location - features to distinguish sequence from different origins

What do you need for QTL analysis?

• (i) A large population of individuals that you can score for phenotypes and genotypes: Recombinant Inbred Lines (RILs)

• (iv) A way to compare identify which markers from each parent have been inherited by the progeny

• (ii) A map of the genome to find out where you are (find out which chromosome the QTL is on)

F1 =Heterozygous

at all loci

(i) A large population of mapping Recombinant Inbred Lines

A Bx

F2 =Heterozygousat some loci

Parents =Homozygous

crossing-over(recombination)

x

F7 RILs =Homozygous

at all loci& heterogeneous

x5Many different individuals are obtained & separately

selfed to develop RILs

• Visible phenotypes or molecular markers (DNA sequence differences)

(ii) Markers to enable identification of which parental genome each part of the chromosomes of the progeny have come from

parent A

parent Bparent A

parent B

Parent AChr 1

(iii) A map of the genome: anchor the markers

Different chromosomes Molecular markers = features of the DNA sequence

Parent AChr 2

Markers differ between parents (natural variants)

Parent AChr 1

Parent BChr 1

Different species variants single nucleotide polymorphisms

GAATTC GATTTC

(iv) You can distinguish these sequence differences using molecular techniques = molecular markers

• Restriction enzymes e.g. EcoRI cut DNA only at a specific recognition sequence

• Compare restriction patterns:

Parent A Parent B........GAATTC.......GAATTC.......GAATTC....... ........GAATTC.......GATTTC.......GAATTC.......

........GAATTC.......GAATTC.......GAATTC....... ........GAATTC.......GATTTC.......GAATTC.......

Second generation (F2)from selfing F1:

First generation (F1)

There are many types of molecular markers

• Restriction Fragment Length Polymorphisms (RFLPs)

• Simple Sequence Length Polymorphisms (SSLPs)

• Cleaved Amplified Polymorphic Sequences (CAPS)

• Microsatellites (repeated sequences of 1-6 bases)

• Essentially, all of these are methods with which to detect sequence differences that have occured between two variants of a species

• They mostly differentiate single nucleotide polymorphisms (snps)






Lecture objectives

Need to know the linkage order: making a genetic map

There are two types of maps:

• Physical map: lays out the sequence information and annotates it: promoters, genes etc.

• 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)

a A

B b

a A

B b

Rf = 0.5 (50%) = no linkage

• Are loci A and B linked (on same chromosome) or unlinked (different chromosomes)?

Genetic linkage is related to recombination frequency

Little recombinationso Rf = small= tight linkage

a AB b

aB, Ab, ab, and ABin equal proportions

Only aB and Ab

Some recombinationso Rf = medium

= quantifiable linkage

a AB b

More aB, Abthan ab, AB

More recombinationso Rf = high ( <0.5 )

= weak linkage

a A

B b

aB, Ab, ab, and ABin similar proportions

Rf = recombinationfrequency

Map distances and genetic linkage

• 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 **

• To make a map you need to score many markers in many individuals

• Recombination frequency of 0.01 (1%) = a genetic map unit of 1 cM

• Recombination events occur randomly, once or twice per chromosome

a AB b

• Assumes that linkage is the only cause of non-independence between markers and that segregation is Mendelian

• Likelihood ODds ratio: likelihood of the observed linkage

• The higher the LOD score, the more closely linked the markers are

Determining map order

• Traditionally done by hand using e.g. the Chi-squared statistic to test for goodness of fit for the observed segregation ratios between markers

• Data on the presence/absence of 100s of markers in (F7) progeny population• Then you can use statistics to work out the marker order

• 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!!

• That’s a lot of Chi-squared tests!• So we use mapping software e.g. Mapmaker, JoinMap

a AB b

Determining map order

A a

B b

• Recombination fraction = n recombinant gametes total

• Haldane mapping function adjusts map distance to account for double crossovers that go undetected

• Kosambi mapping function also adjusts for crossover interference i.e. a crossover reduces the probability of a second crossover nearby

C c• Map distance ≈ (RAB + RAC - 2RABRBC) x 100 cM

• 2RABRAC is negligible for <10cM

These should theoretically correspond to chromosomes, but if...

• Chromosomes very long

• Recombination frequency very high

• Mapping populations are not large enough

...one chromosome can statistically “break” into several linkage groups

• Also, centromeres and heterochromatin have supressed recombination

Linkage groups are the basis of genetic maps

A genetic linkage map for broccoli1 2 3 4 5 6 7 8 9

map unitscM

• Recombination frequency of 0.01 (1%) = a genetic map unit of 1 cM

Download - CH927 Quantitative Genomics What is the genetic basis of ...By the end of this lecture you should be able to explain: • Quantitative genetics: homozygotes, heterozygotes and inheritance

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