Autozygosity Mapping and Linkage Analysis Flashcards
What is a Mendelian trait?
A measurable characteristic of an organism that is inherited following the patterns determined by Gregor Mendel. A disease can be a Mendelian trait, and many monogenic diseases are examples of this.
Describe 7 characteristics of an ideal genetic marker.
- Polymorphic
- Randomly distributed across the genome
- Known location in genome
- Frequent in genome
- Frequent in population - preferably minor allele freq >10%
- Stable with time
- Easy to assay (genotype)
Describe the 3 most common genetic markers and their uses.
Microsatellites
- Short repeats of <10bp, multi-allelic, random, frequent
- Linkage and autozygosity mapping
Minisatellites
- Large repeats of 500>bp, multi-allelic, non-random
- DNA fingerprinting
SNPs
- Single-base, bi-allelic, random, frequent
- Linkage and assocation/autozygosity mapping
Explain the approach of linkage analysis and when it is used.
- Gene mapping.
- Using an observed locus (marker) to draw inferences about an unobserved locus (disease gene).
- Family-based design - from few large families to many small nuclear or sibpairs. The bigger the family the more information, and combining multiple families useful.
Explain what is meant by genetic linkage.
- The tendency for alleles at neighbouring loci to segregate together at meiosis is the phenomenon of genetic linkage.
- Therefore to be linked, two loci must lie very close together.
- Alleles at linked loci are known as a haplotype. Haplotypes mark chromosomal segments which can be tracked through pedigrees and populations.
- Cross-overs are more likely to occur between loci separated by some distance than those close together.
What is recombination frequency?
Recombination frequency = ϴ (theta) = RF: the proportion of recombinants in the total number of offspring. Thus, RF is how frequently recombination occurs in a family.
Explain how genetic distance is different from physical distance, and how it is quantified.
- The genetic distance over which 1 cross-over occurs in every gamete is a Morgan.
- A centi-Morgan (cM) or map unit is the distance over which one recombination is seen in every 100 gametes, i.e. RF = 1%.
- Low RF shows linkage between two loci.
- RF defines genetic distance, and a genetic map is the expected distribution of genes based on recombination frequencies.
- Since there are recombination ‘cool-spots’ and ‘hot-spots’ in the genome, the actual physical distance may be greater or less than that predicted by the genetic distance.
Explain what is meant by linkage disequilibrium.
- Describes the relationship between alleles of two loci.
- If alleles at each loci are in close proximity they can be in LD.
- Segments of DNA that segregate together are said to be linked. If the degree of linkage exceeds that expected by chance, the regions are said to be in linkage disequilibrium.
- In LD, RF tends towards 0 as the alleles are in close proximity and recombination is rare.
Explain the difference between identity by state (IBS) and identity by descent (IBD).
- IBS - two individuals share an identical base sequence in a particular DNA segment.
- IBD - an IBS segment shared by two individuals that has been inherited from a common ancestor without recombination.
- DNA segments that are IBD are by definition IBS, but IBS segments are not necessarily IBD.
How are microsatellites genotyped?
- Microsatellites have unique flanking sequences.
- Design primers specific to the flanking sequences.
- Isolate DNA from individuals to be studied.
- PCR amplification.
- Gel electrophoresis.
A genetic distance of 1 cM is roughly equivalent to what physical distance?
1 cM ~ 1 Mb = 1,000,000 base pairs
Explain what is meant by a LOD score and how it is calculated.
- LOD score = log of the odds
- LOD = log10[L(ϴ)/L(0.5)]
- Essentially a likelihood ratio
- Log(odds that loci are linked at ϴ<0.5)/(odds that loci are unlinked, ϴ~0.5)
- I.e. the statistical probability that two loci are genetically linked and not co-inherited by chance.
What LOD score thresholds are considered to provide evidence of linkage and non-linkage?
- LOD >3 is evidence of linkage.
- LOD between 2.9 and -1.9 indicates possible linkage.
- LOD <-2 evidence of no linkage.
Explain how linkage analysis could be used to identify a region of interest containing a disease gene.
- Genotype many genetic markers across the genome - e.g. microsatellites.
- Generate linkage maps, potentially for multiple chromosomes.
- Linkage maps will show LOD scores for all of the markers.
- A linkage peak of LOD scores >3 will indicate the likely region containing the disease gene.
Describe some of the problems with linkage analysis.
Linkage analysis is compromised when there is deviation from classical Mendelian inheritance:
- Reduced penetrance - genotype doesn’t always result in phenotype
- Phenocopy rate - non-genetic (environmental) influences resulting in the same phenotype
- Locus heterogeneity - same disease can be caused by mutations in different genes - relevant to linkage analysis involving multiple families
- Variable expressivity - different presentation/severity of disease can result in individuals being considered affected/unaffected
