1-31 Linkage and Recombination (Clinical Applications)

Question 1

What is the difference between direct and indirect linkage analysis? Why would anyone use indirect linkage analysis?

Answer 1

Direct linkage analysis investigates the direct source of a disease-causing mutation; indirect linkage analysis uses linked markers. Reasons to use indirect linkage analysis include:

• Too many mutations
• Gene is too large
• The disease-causing gene, although mapped, is unknown
• Often impractical and/or too costly to find the specific disease-causing mutation

Question 2

What are some potential issues with indirect linkage analysis?

Answer 2

• Need correct disease diagnosis and no locus heterogeneity (or first have to use linkage to establish which locus is involved)
• DNA from critical family members will be needed and their cooperation required
• Paternity needs to be as stated!
• Need to find a marker that is “informative”
• If using extragenic markers there maybe an error rate associated with recombination; thus intragenic markers are better than extragenic because recombination is usually then not an issue

Question 3

What are polymorphisms and rare genetic variants?

Answer 3

Polymorphisms: alleles/mutations that occur at a frequency of 1% or greater; i.e., 2% or more of individuals will carry at least one autosomal copy (be heterozygous OR homozygous)

Rare genetic variants: alleles that occur at a frequency of less than 1%

Question 4

What is recombination?

Answer 4

The formation of new combinations of linked genes by crossing over between loci. This process occurs via reciprocal exchange of segments between chromatids of homologous chromosomes at prophase of the first meiotic division.

The distance between genes is measured in centimorgans (cM). 1cM = 1% chance of crossover. The greater the distance, the more likely the chance of crossover.

Question 5

What is a haplotype?

Answer 5

Genes/loci located close enough together on the same segment of chromosome that they tend to travel together through meiosis.

Question 6

In the following image, recombination fraction (solid line) and map distance (dotted line) are nearly equal, with 1 cM = 0.01 recombination, for values of genetic distance below 10 cM. Why do they diverge?

Answer 6

Recombination fraction and map distance diverge because of double crossovers as the distance between the markers increases. The recombination fraction approaches a maximum of 0.5 no matter how far apart loci are; the genetic distance increases proportionally to the distance between loci.

Question 7

What is linkage disequilibrium?

Answer 7

The tendency of specific combinations of alleles at 2+ linked loci to occur together (in coupling) on the same chromosome more frequently than would be expected by chance. This phenomenon represents a deviation from Mendel’s second law (independent assortment).

D = (genotype frequencies) - (product of the allele frequencies)

Linkage disequilibrium decays with recombination distance and time (generations), but it can indicate that loci are close and can be used to indicate that gene mapping is closing in on the target gene e.g. km19 in mapping CFTR gene.

Question 8

What are the 3 kinds of “labels” used in mutation testing?

Answer 8

1. Linked extragenic markers (indirect): like a label attached to a suitcase by a length of string. Problem: the marker (label) can become detached from the gene (suitcase)
2. Intragenic markers (indirect): like a label on a suitcase. Much less likely that the marker and gene will become separated, unless . . . Problem: DMD intragenic markers have a significant recombination rate with the disease-causing mutation.
3. Disease-causing mutations (direct): like the bomb inside the suitcase. You don’t need the family to look for the mutation–it’s there.

Question 9

Which markers are preferable: flanking or single side?

Answer 9

Flanking markers are preferable, because they would require a much rarer double crossover to cause error.

(They would, however, be incorrect 1% of time if relying on markers 1 cM away on same side.)

Question 10

What are coupling and repulsion?

Answer 10

Coupling/cis: 2 alleles located on the same chromosome and inherited from the same parent

Repulsion/trans: 2 alleles located on different homologous chromosomes and inherited from different parents

Question 11

What is phase, and how can we know it?

Answer 11

When mutations are known to be on the same or different homologous chromosomes, i.e., known to be in coupling or repulsion, then phase is known.

Typically, phase is determined by coming down a pedigree.

Question 12

What is a lod score?

Answer 12

A lod score (Z) is a statistical method that tests genetic marker data in families to determine whether two loci are linked. The lod score is the logarithm to base 10 of the odds in favor of linkage.

• A lod of 3 (1000:1 in favor) in an autosome or 2 in an X-linked is taken as proof of linkage
• A lod of -2 (100:1 against) is proof that the loci are unlinked
• Threshold for a genome-wide level allowing for multiple markers is 3.3; for non-Mendelian characters, any score below 5 should be regarded as provisional