Unit II- Linkage and Recombination (Clinical Applications)

Question 1

Indirect Testing with markers

Answer 1

• whenever it is not possible to test for disease-causing mutations either because there are too many mutations or because the disease causing gene, although mapped, is not known, it may still be possible to use linked markers to perform carrier testing and/or prenatal diagnosis
• when a genetic disease has a large gene and many mutations, it often impractical and/or too costly to find the disease -causing mutation
• an alternative is to perform an indirect test using a linkage analysis with linked markers

Question 2

Disadvantage to indirect testing with markers

Answer 2

• need correct disease diagnosis and no locus heterogeneity (or use linkage to establish which locus involved)
• DNA from critical family members will be needed and their cooperation required. Paternity needs to be stated
• need to find a marker that is “informative” i.e. need to flag the “bad” gene to answer the clinical question being asked
• 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

Polymorphisms: a way of flagging chromosomes

Answer 3

• need to flag the chromosomes so as to follow specific chromosomal locations through meiosis
• examples:
• single nucleotide polymorphisms
• microsatellites (variable number of tandem repeats VNTRs)

Question 4

Mutation

Answer 4

-is a permanent heritable change in genomic DNA sequence, often used incorrectly to indicate a “bad” change

Question 5

Polymorphism

Answer 5

-a sequence variant found at a frequently of at least 1% (at least 2% of people are heterozygotes), often incorrectly used to mean benign

Question 6

Rare genetic variant

Answer 6

-is a mutation with a frequency of less than 1%

Question 7

Family versus marrying in relatives

Answer 7

-disease allele is linked to 2 in this family, it does not mean it will always be linked to the 2 in other families or that everyone with a 2 also has the disease-causing mutation

Question 8

Recombination

Answer 8

• the formation of new combinations of linked genes by crossing over between loci
• there are on average three crossovers per chromosome per generation

Question 9

Linkage

Answer 9

genes close enough together on the same chromosome have a tendency to be transmitted together through meiosis more often than expected by chance
-linked genes form haplotypes

Question 10

Haplotype

Answer 10

• a group of alleles from closely linked loci, usually inherited as a unit
  e. g. a set of restriction fragment length polymorphisms closely linked to one another and to a gene of interest
  e. g. the D and M loci in B panal but not A panal on the next slide

Question 11

Crossing over

Answer 11

• the reciprocal exchange of segments between chromatids of homologous chromosomes
• mechanism of recombination

Question 12

Recombination Distance

Answer 12

• 1 centiMorgan is a 1% change of recombination between loci as the chromosome is passed from parent to child per generation
• the recombination fraction approaches a maximum of 0.5 no matter how far apart loci are

Question 13

Linkage disequilibrium

Answer 13

• the tendency of specific combinations of alleles at two or more linked loci to occur together on the same chromosome more frequently than would be expected by chance
• the deviation from Mendel’s second law of Independent Assortment
• is the difference between the genotype frequencies and the product of the allele frequencies
• decays with recombination distance and time
• 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 14

Linkage disequilibrium in CF

Answer 14

• the delta F508 CF mutation arose once. It arose on a chromosome 7 with the 2.1kb Taq I allele at xv-2c and the and the 6.6 kb Pst I allele at km19 which are marker loci about 60 kb from CFTR
• over the succeeding generations there have been few crossover events between these marker loci and the CFTR gene such that the Delta F508 mutation is still usually found (>90%) with these alleles at these loci i.e. on this background

Question 15

Three classes of mutations

Answer 15

• linked extragenic markers- attached to suitcase by length of string
• intragenic markers (label on suitcase)
• disease-causing mutation (see inside suitcase)

Question 16

DMD gene

Answer 16

• The dystrophin gene is the exception
• DMD is where the intragenic markers may still have a significant recombination rate with disease-causing mutation
• flanking markers are more useful since it takes a double cross over, rather than a single cross over to cause an error

Question 17

Coupling and repulsion

Answer 17

• alleles at different loci are said to be in coupling when on the same chromosome i.e. in cis
• alleles at different loci are said to be in repulsion when on opposite chromosomes i.e. in trans

Question 18

Phase

Answer 18

• when mutations are known to be on the same or different homologous chromosome i.e. known to be in coupling or repulsion, then phase is known
• when it is not known which mutation is on which homologous chromosome i.e. whether the alleles at different loci are in coupling or repulsion, then phase is unknown

Question 19

Lod Score

Answer 19

• 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
• by concention a lod of 3 in autosome or 2 in an X-linked is taken as proof of linkage, and a lod score of -2 as proof that the loci are unlinked
• threshold for a genome-wide level allowing for multiple markers is 3.3 while for Non-Mendelian characters, any score below 5 should be regarded as provisional

-series of likelihood ratio are calculated at various possible recombination distances theta and plottoed in a table or graph