Association analysis

Question 1

What is genetic association?

Answer 1

presence of a variant allele at a higher frequency in unrelated subjects with a particular disease of interest (cases) compared to those that do not have the disease (controls)

Question 2

What are cases?

Answer 2

subjects with the disease of interest

Question 3

What are the controls in association analysis?

Answer 3

must be identical to out cases APART from not having the disease

e.g. same age, sex, ethnicity, location etc.

Question 4

What is the Difference in variant frequency between cases and controls

Answer 4

in cases, the gene variant is at a higher frequency than in the controls and is associated with the disease

Question 5

What confirms the strength of association between a gene variant and the disease?

Answer 5

Statistics e.g. p-value

Question 6

What do quality case control genetic studies require?

Answer 6

• large numbers of well defined cases (1000s)
• equal numbers of matched controls
• reliable genotyping technology (SNP array)
• standard statistical analysis
• positive associations should be replicated

Question 7

What allows us to capture genetic diversity?

Answer 7

The use of genetic markers

Question 8

Give some Features of an ideal genetic marker (e.g. SNP).

Answer 8

• polymorphic
• randomly distributed across the genome
• fixed location in genome
• frequent in the genome
• frequent in the population
• stable with time
• easy to assay (genotype)

Question 9

How are SNPs generated?

Answer 9

though mismatch repair during DNA replication

Question 10

What are the possible locations for SNPs?

Answer 10

Gene (coding region)

• no amino acid change (synonymous)
• amino acid change (non-synonymous)
• new stop codon (nonsense)

Gene (non-coding region)

• promoter: mRNA and protein changed
• terminator: mRNA and protein changed
• splice site: altered mRNAm altered protein

Intergenic Region
-98% of SNPs in this region

Question 11

What is dbSNP?

Answer 11

online database of SNPs and multiple small-scale variations that include insertions/deletions, microsatellites and non-polymorphic variants

Question 12

What is the Minor allele frequency (MAF)?

Answer 12

the frequency of the less common variant in a population

SNP will have two alleles:

• major allele
• minor allele

Question 13

What is the sum of the major and minor allele frequencies?

Answer 13

Major Allele Frequency + Minor Allele Frequency= 1

Question 14

What are the Genome Wide Association Studies (GWAS)?

Answer 14

Studies of variations in the entire human genome to identify associations between variations in genes and particular behaviours, traits, or disorders.

SNP markers are used across the whole genome, and these are genotyped using SNP microarrays.

We look for association between disease and each marker by doing a chi-square test

Question 15

Explain the steps of GWAS.

Answer 15

Obtain DNA from people with disease of interest (cases) and unaffected controls

Run each DNA sample on a SNP chip to measure genotypes at 300,000-1,000,000 SNPs in cases and controls

Identify SNPs where one allele is significantly more common in cases than controls
-the SNP is associated with disease

Question 16

How is GWAS data presented?

Answer 16

on a Manhattan plot:
The end result from GWAS that shows the score for each SNP marker tested in the study, across each chromosome in the genome - a peak indicates that the location of that particular SNP may be very close to a disease-related locus - low p-value = high peak –> reject null hypothesis for that SNP - shows evidence of correlation of each marker tested with a disease phenotype across the genotype

*highest point is MOST significant SNP with disease state

X-axis is position of SNP on chromosome

Y-axis is -log10(p-value) of the association between each marker and the disease, calculated using a chi-squared test

Question 17

Why are chromosomes more solid at the bottom of the Manhattan plot?

Answer 17

because the vast majority of SNPs are not associated with disease

*associations are further up the graph

Question 18

What is the Wellcome Trust Case Control Consortium?

Answer 18

genome-wide association study of 14,000 cases of 7 common diseases and 3,000 shared controls

• looks for specific genes
• look at p-value (the smaller means stronger association)
  ex) LDLR, SORT1 for lipids and CAD

Question 19

What does the peak on a Manhattan plot show?

Answer 19

The peak does not identify the gene causing the disease, instead it only identifies the genomic REGION associated with the disease and this is usually very small (<100kb)

-if you zoom in on an SNP with strong disease association (high up in plot), you will find lots of other associated SNPs in the adjacent gene and therefore more work has to be done to work out which SNP is correct and where the actual disease-causing variant is

Question 20

Give some Disadvantage of GWAS.

Answer 20

Difficult to do very large association studies (>1K cases), and therefore meta-analysis of GWAS is done to combine statistical results from multiple smaller studies

Pre-experiment: consortium
Post-experiment: meta analysis

Question 21

Give the Medical Complications of Obesity.

Answer 21

Pulmonary Disease
Non-alcoholic fatty liver disease
Gall bladder disease
Osteoarthritis
Skin
Gout
Phlebitis
Cataracts
Coronary Heart Disease
Cancer
Severe Pancreatitis
Stroke
Idiopathic intracranial hypertension
Gynaecological Abnormalities

Question 22

Give some of the problems of GWAS.

Answer 22

Expensive

requires large sample size,

only explains a small part of the disease phenotype, meaning their contribution to the genetic component of the disease is estimated to be low (<5%), could be because:

• many common SNPs of small effect
• rare SNPs
• don’t look at copy number variation or epigenetic variation

Question 23

How are genes associated with obesity in GWAS?

Answer 23

Obesity is strongly genetic

Though GWAS we can clearly see genes associated with obesity