Gene Mapping and Disease Gene Cloning

Question 1

Linkage analysis

Answer 1

ID rare and highly penetrant defects in single genes that cause Mendelian conditions. Physical proximity of genetic loci by examining the frequency of crossing over or the recombination fraction between genetic markers in a series of families with affected individuals. Include polymorphic microsattelite repeats (di/tetra repeats), SNPs, RFLP. The closer the two gametes are, the less likely crossing over will occur. Must analyze families to get the disease gene.

Question 2

Recombination Fraction

Answer 2

Proportion of the off spring who are recombination. Proportion of meiosis that results in recombinant. Denotes the frequency of crossing over between two loci during mitosis. Genetic distance is measured in cM centimorgans. A recombination fraction of 1% is equal to a genetic distance of 1 cM, which in turn is equivalent to a physical distance of approximately 1 million nucleotides between loci. Genetic distance is not always equal to physical distance.

Question 3

Logarithm of the odds (LOD) score

Answer 3

Z score. A measurement of the likelihood that two loci are linked at a given recombination fraction (denoted theta) is divided by the likelihood that two loci are not linked at theta is .5. Values range from 0-.5 for theta. The value of theta of which LOD is highest is considered the most likely genetic distance between the two loci. An LOD greater than 3 is considered evidence for linkage. An LOD score of -2 is considered evidence against linkage. Scores in between is inconclusive.

Question 4

Recombination fraction

Answer 4

Ranges from 0% to 50%. 50% simply represent independent assortment while 0% is linkage?

Question 5

Association Studies

Answer 5

Co-occurance of two traits more often than expected by chance. Requires populations study rather than family study. Susceptibility genes to common complex diseases such as diabetes. Relative risk or odds ratio in a case control study.

Question 6

Linkage Disequilibrium

Answer 6

Special form of association. A set of loci closely linked on a chromosome and co-segregate with a disease phenotype in a population. Assessed by genome-wide association studies. Test frequencies of a set of loci (SNPs) in case and control to search for loci that co-segregate with a trait. A strong linkage disequilibrium suggests that the gene itself or those in close proximity are susceptibility genes.

Question 7

Physical Mapping

Answer 7

Analysis of chromosomal abnormalities. Characterizations of deletions or translocation breakpoints led to discovery of several disease genes.

Question 8

Functional Mapping

Answer 8

Genes mapped based on knowledge of the gene product. For example, first derived from its amino acid residues, then mapped out after.

Question 9

Next-Gen Sequencing

Answer 9

Limiting factors for finding disease genes include smaller number of cases for family studies, reduced penetrance, locus heterogeneity, and diminished reproductive fitness. NGS is a powerful tool in discovery of disease genes that are intractable by the conventional approaches. Whole exome sequencing WES. Whole genome sequencing WGS. Do not require whole families but bioinformatics remains a challenge.

Question 10

Disease Gene Validation

Answer 10

Candidate genes require further studies. Bioinformatics analysis of gene sequences on protein function. ID of disease specific variants that co-segregate with the patients but not with healthy controls. Biochemistry function analysis. Model organisms.

Question 11

Positional Mapping

Answer 11

Employs different approaches to identify a candidate region on a chromosome in which candidate genes may reside, followed by testing these genes to identify the gene of interest. This strategy has played an important role in disease gene discovery.