Transcript Genetic Markers, Linkage and Association Genetics 202

APPROACHING THE GENOME GENETIC MARKERS, LINKAGE AND
ASSOCIATION
GENETICS 202
Jon Bernstein
Department of Pediatrics
October 8, 2015
Session Goals

Develop an understanding of genetic markers and what they can be
used for

Learn how structural variants in chromosomes have aided the
identification of genetic loci associated with various diseases

Understand in conceptual terms how a linkage study is performed

Understand in conceptual terms how an association study is
performed and how they are related to SNP based risk assessment
Lecture Outline

Mapping genes by the stumble upon method
◦ Chromosomal anomalies and CNVs in gene discovery

Introduction to genetic markers
◦ Linkage studies
◦ Association studies
Current knowledge of gene – disease
relationships
Matching genotypes and phenotypes

It must be in there
somewhere….
Stanford Cytogenetics Lab
What do methods for matching genotypes and
phenotypes have in common?

Based on statistical evidence
◦ Core question is: Is a genotype occurring with a phenotype
more than would be expected by chance
 Can be looked at in families
 Unrelated individuals
The story of CHARGE
Sanlaville and Verloes, EJHG, 2007
www.chargesyndrome.org
Nature Genetics, September, 2004, PMID: 15300250
Microduplications and deletions as CNVs

Copy number variant (CNV)
◦ A gain or loss of a contiguous block of DNA
 1Kb to several Mb
◦ Referred to as a “variant” as this does not imply
pathogenicity
CNVs are a part of normal genetic variation
If the human genome project was completed in
2003, why are widespread CNVs reported later?
What if you cannot find a rare cytogenetic
abnormality ?

Mapping traits to genes by
◦ Linkage studies
◦ Association studies
Mapping DNA, Mapping traits to DNA

In general, the closer two loci are together the less
likely it is that they will be separated by a random
break in the chromosome they are on.
A
B
C
Fig 9.2 Crossing over between paired homologous chromosomes in the first division of meiosis produces recombination between genetic loci
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
The effect of multiple meioses on a chromosome
http://hapmap.ncbi.nlm.nih.gov/originhaplotype.html.en
Genetic Markers

Variable or polymorphic DNA
elements that are readily assessed
by molecular biology methods

Facilitate the differentiation of one
copy of a locus from the other
http://hapmap.ncbi.nlm.nih.gov/originhaplotype.html.en
Common types of DNA polymorphism used as markers
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Paternity testing using genetic markers
6,9.3
7,9
6,7
Figure 1 : Paternity testing using microsatellite markers.
This test includes samples from the mother (top row), the child (middle row), and the
alleged father (bottom row). The maternal marker that has been passed to the child is
6. This means that the other marker present for the child (7) must have been inherited
from the father. The alleged father matches the child, since one of his markers is
indeed 7.
Adams, J. (2008) Paternity testing: blood types and DNA. Nature Education 1(1)
UPD Studies and genetic markers
Non-paternity versus UPD
Looking for linkage between a marker and a disease
locus
Trait locus
2
•Informative vs Uninformative
Markers (Can we tell which
marker segregated with the
trait)
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
•Recombinant and nonrecombinant genotypes
2
Marker
Trait locus
4
2
Marker
Looking for a gene linked to dyschromatosis on
chromosome 1
Determine the odds/relative
likelihood of the pedigree
(affected/unaffected status)
given the genotyping results in
each family.
 Results are expressed as a log of
odds or LOD score.
◦ A score of one means ten
times more likely
◦ Cutoff score of three
typically used (1000 times
more likely)

Zhang et al., Journal of Investigative Dermatology, 2003, (120) 776-180.
Fig 9.6 Pedigrees of the three families used by Miyamura et al. (2003) to map the dyschromatosis gene (Part 1)
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Linkage analysis results

High LOD scores at
multiple markers in the
center of the region of
interest on chromosome
1q.
Fig 9.6 Pedigrees of the three families used by Miyamura et al. (2003) to map the dyschromatosis gene (Part 1)
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Fig 9.7 Haplotypes of seven individuals from the three pedigrees in Figure 9.6
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Using linkage in clinical diagnostics

Used when mutation not
identified or gene
unknown at a locus

Caveats
◦ Recombination can occur
◦ Family members must be
phenotyped correctly
www.genereviews.org
What types of diseases is linkage best for
Show Mendelian
inheritance
 High heritability
 High penetrance
 Genetically
homogenous

Manolio et al., Nature, October, 2009
What types of diseases are best studied by
GWAS?
 Common
 Relatively high
heritability
 Caused by variants
of moderate effect
Manolio et al., Nature, October, 2009
Association Studies – An Alternative to linkage

Look for a statistical association between a genetic
variant and a trait or disease.
◦ Can look at genetic variants at a few loci or thousands at a
time.
Genome Wide Association Studies (GWAS)

Often done as case control studies in which
associations between thousands of genetic loci and a
traits or traits are tested for simultaneously.

Based on the hypothesis that ancestral common
variants are associated with common disease.
Where do common genetic variants
come from?
Data from census.gov, graph from Wikipedia entry on World Population
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Linkage Disequilibrium (LD) and Haplotypes in
Association Studies
LD: Nonrandom
association of alleles
at 2 or more loci
Haplotype: A group of
adjacent genetic
elements of a
chromosome that is
transmitted together
http://hapmap.ncbi.nlm.nih.gov/originhaplotype.html.en
SNP Genotyping – Rapid Assessment of
Many Markers
•Routinely done on
specialized DNA arrays
•500K – 1M+ SNPs on a
single chip
LaFramboise T Nucl. Acids Res. 2009;37:4181-4193
Haplotype blocks

LD blocks vary in size
across the genome, but
are estimated to be on
the order of 10-50kb
ADRA1A locus at 8p21.2
Buzas B et al. Mol Genet Genomics. 2004 Dec;272(5):519-29. PMID: 15503142
P<5×10−8
Manolio, NEJM, July 2010, PMID: 20647212
Fig 13.8 An overview of genetic susceptibility factors identified by genome-wide association studies (Part 1)
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
Fig 13.8 An overview of genetic susceptibility factors identified by genome-wide association studies (Part 2 – key)
New Clinical Genetics 2e
Andrew Read and Dian Donnai
ISBN: 9781904842804
© Scion Publishing Ltd, 2011
No association
Pitfalls in GWAS

False negatives
◦ Inaccurate phenotyping
 Miscategorization of cases and
controls
◦ Other potential causes
 Genetic heterogeneity
 The presence of many genetic contributors makes
each association harder to detect

Affected
Unaffected
Variant
5
45
No Variant
5
45
Complete association
Affected
Unaffected
Variant
10
0
No Variant
0
90
Random assignment of affected
False Positives
◦ Population stratification
Affected
Unaffected
Variant
5
5
No Variant
45
45
Sample GWAS Results
Wellcome Trust Case Control Consortium. Genome-wide association study of 14,000 cases of seven
common diseases and 3,000 shared controls. Nature. 2007 Jun 7;447(7145):661-78. PMID: 17554300
Controls vs controls
Healthy individuals of different
ancestry (multiple groups)
GWAS Results for Personal Risk Assessment

Identification of SNPs associated with disease can be
used to assess the risk of individuals who were not in the
original study.
GWAS Results for Personal Risk Assessment
Chen, Rui et al. Cell , Volume 148 , Issue 6 , 1293 – 1307, PMID: 22424236
Ancestry informative markers
1384 individuals, 1200 markers (729 microsatellite and 471 insertion/deletion)
Rosenberg et al., PLOS Genetics, December 2006, PMID: 17194221
The “Heritability Gap”
Nature, October, 2009
Potential explanations of the heritability gap




Rare variants play a significant role in common diseases
There are numerous genes each with small effect in the
etiology of common diseases
◦ Genetic heterogeneity
◦ Small effect size
There are significant gene-gene and gene-environment
interactions
Heritability has been overestimated
New Methods for Discovering Disease Gene
Associations



Structural variant (CNV) based association studies
Rare single nucleotide variant association studies
Enable the detection and study of rare variants not
present in ancestral populations
CNVs as an Example of Rare Variants and
Common Disease
Nature 461, 747-753 (8 October 2009)
Exome based rare variant discovery – Kabuki
syndrome and MLL2
Ng SB et al. Exome sequencing identifies MLL2 mutations as a cause of Kabuki
syndrome. Nat Genet. 2010. PMID: 20711175
Lecture Summary


A number of strategies have been used to develop our
current knowledge about relationships between genes and
disease
◦ Structural chromosome variants
◦ Linkage
◦ Association
Genetic markers are variable or polymorphic elements within
DNA that allow the differentiation of corresponding regions
on homologous chromosomes
Review Question

Both linkage studies and GWAS studies are likely to
produce false negative results when
◦ A)There is high heritability
◦ B) There is genetic heterogeneity
◦ C) The disease is common
◦ D) There is population stratification
Review Question

To be most useful in conducting linkage and association
studies, DNA markers should ideally be?
◦ A) Homozygous
◦ B) Highly polymorphic
◦ C) Very widely spaced in the genome
◦ D) Very rare in the population