Mapping Mendelian Disease

Question 1

As a recap what are the different classifications of genetic diseases?

Answer 1

• MENDELIAN/MONOGENIC - disease caused by a single gene e.g PKD (polycystic kidney disease)
• NON-MENDELIAN/ POLYGENIC - disease or traits caused by the impact of many different genes, each onlt has small individual impact on the final condition e.g psoriasis
• MULTIFACTORAL - disease or traits resulting from an interaction between multiple genes and often multiple environmental factors e.g heart disease

Question 2

How do we study mendelian diseases?

Answer 2

Gene identification by gene mapping

• Homozygosity mapping
• Linkage analysis
• GWAS

Question 3

How do we find disease-causing mutations?

How do we prove they cause a disease?

Answer 3

• By sequencing
• Using in silico, in vitro and in vivo tools

Question 4

Recap of the principles of genetic linkage

Answer 4

Genetic linkage is the tendency for alleles at neighbouring loci to segregate together at meiosis

A haplotype defines alleles at linked loci. Haplotypes mark chromosomal segments which can be tracked through pedigrees and populations.

Cross-overs are more likely to occur between loci at some distance

Question 5

How do we use the principles of genetic linkage to identify disease causing genes?

Answer 5

• If a marker is linked e.g M3 and M4 the same marker alleles will be inherited by two affected relatives more than expected by chance
• If the marker and diseased locus are unlinked (M5-M8), the affected relatives are less likely to inherit the same marker alleles
  
  • The markers are usually SNP markers chosen from databases and we know exactly where they are.
    
    • You will then zoom into the diseased gene region and find out what genes lie in that region and which may be the disease causing gene

Question 6

Recap what linkage analysis is

Answer 6

Linkage analysis is a method used to map the location of a diseased gene in a genome

It is carried out using an observed locus (marker) to draw interferences about an unobserved locus (diseased gene).

Our goal is to find the genomic regions linked to disease

Question 7

Summarise the process of linkage analysis

Answer 7

1. Take a pedigree
2. Use some kind to tool to generate genotyping data for your pedigree (e.g genotyping SNP array)
3. Graph the physical and genetic distribution of markers on a genotyping array
4. Run linkage programme (choose to run it in a parametric or non-parametric way)
5. Figure out LOD score

Question 8

What is the difference between running a linkage programme in a parametric and non-parametric way?

Answer 8

PARAMETRIC = imposes rules about inheritance and disease frequency

NON-PARAMETRIC

• doesn’t assume anything about inheritance patterns
• In some graphs there are peaks and in some there is linkage (this gives us a rough idea of what could happen if you start applying the model)
• There are no rules imposed in NPL, looks for IBD (identity by descent)

Question 9

Explain LOD scoring

Answer 9

LOD scores asseses whether the linkage between two loci is due to chance or not

The higher the LOD score, the higher the likelihood of linkage

(LOD scores are additive - different famillies linked to the same disease locus will increase the LOD score)

• LOD score greater than 3.0 is taken as siginificant
• LOD score between -2 and 3 are inconclusive
• LOD score below -2.0 shows significant non-linkage

Question 10

What is the role of the lymphatic system?

Answer 10

Fluid homeostasis

Immune function

Fatty acid transport

Question 11

Describe primary lymphoedema

Answer 11

Here the lymphatic system has not developed properly or isnt functioning properly

Is a chronic oedema is caused by a developmental abnormality of the lymphatic system- affecting 1 in 6,000 people

• Phenotypes vary
  
  • age of onset
  • site
  • inheritance patterns
  • associated features
  • genetic causes

Question 12

What is the current treatment available for lymphoedema?

Answer 12

• Manual lymph drain massage (MLD)
• Bandaging

Question 13

What is generalised lymphatic dysplasia, Hennekam Syndrome?

Answer 13

• generalised as it covers the whole body
• Antenatal hydrops with ascites and pleural effusions
• Oedematous at birth
• Intestinal lymphangiectasia (swelling of intestines)
• Peripheral lymphoedema; arms, legs, face
• Mild developmental delay

Question 14

Using the example of lymphoedema, how would you go about finding the gene responsible for the disease?

Answer 14

1. Generate genotyping data - carried out using Genome-Wide Human SNP Array 6.0
2. Run linkage analysis using a autosomal recessive model. The result in this case showed linkage to chromosome 18
3. Next you would find the mutation causing the disease in the candidate gene
4. This would be carried out by designing a primer for each exon of the candidate gene and carry out Sanger Sequencing
5. Final step = proving mutations in the gene are identified as disease causing e.g knock genes into zebra fish and see they are filling up with fluid

Question 15

Describe 4-limb lymphoedema

Answer 15

• Autosomal dominant
• Pubertal/adult onset
• Associated with venous incompetence
• No other abnormalities

Question 16

How do we find disease causing mutations?

Answer 16

• Traditional Sanger sequencing
  
  • Candidate gene screen
• Next generation sequencing (NGS)
  
  • Whole genome sequencing (WGS)
  • Whole exome sequencing (WES) (whole exome sequencing is usually quicker because rare variants that we are looking at are in the coding region which means we do not require the whole genome)