Presentation on theme: "Human Gene Mapping and Disease Gene Identificaton"— Presentation transcript:
1Human Gene Mapping and Disease Gene Identificaton 10ChapterHuman Gene Mapping and Disease Gene IdentificatonPaul CouckeJan HellemansAndy Willaert1
2The genetic landscape of the human genome Chapter 10The genetic landscape of the human genomeMapping human genes by linkage analysisMapping of complex traitsFrom gene mapping to gene identification
3The genetic landscape of the human genome AimsThe genetic landscape of the human genomeIndependent assortement and homologous recombination in meiosisRecombination frequency and map distanceLinkage equilibrium and disequilibriumThe hapMap3
4Mapping human genes by linkage analysis Theory Practise AimsMapping human genes by linkage analysisTheoryPractiseInterprete microsatellite resultsAdd genotypes to pedigreesCreate pedigree and genotype filesCalculate and interprete LOD-scoresDelineate linkage intervalsFocus on Excel because this is the program where you are most likely to get some quick benefits from VBA.4
5Importance of gene mapping : Immediate clinical application as it can be used in prenatal diagnosis,presymptomatic diagnosis and carrier testing.A first step in the identification of a disease gene (positional cloning).An opportunity to characterize the disorder as to the extent for exampleof locus heterogeneity.Makes it possible to characterize the gene itself and the mutationsinvolved resulting in a better understanding of disease pathogenesis.
7Importance of gene mapping : Immediate clinical application as it can be used in prenatal diagnosis,presymptomatic diagnosis and carrier testing.A first step in the identification of a disease gene (positional cloning).An opportunity to characterize the disorder as to the extent for exampleof locus heterogeneity.Makes it possible to characterize the gene itself and the mutationsinvolved resulting in a better understanding of disease pathogenesis.
8The genetic landscape of the human genome I will go slowly over the basics as it is important that you can follow the reasoning behind the different formulas. I apologize for those who are already familiar with this matter.
9recombination in meiosis Each of you know that for each meiosis haploid gametes are formed through homologous recombination between chromosome pairs as explaind in chapter 2. To understand genetic linkage analysis it is important to understand the behavior of chromosomes and genes during meiosis.
10recombination in meiosis Alleles at loci on differentchromosomes assortindependentlyAll of you know that two loci located on different chromosomes assort idependently. In fact 4 different possibilities are generated
11Recombination frequency (theta) Ofcourse it is different when the two loci are located on the same chromosome. In this case there are 3 possibilities.The amount of recombinations between two loci is therefore ameasure for the distance between these two loci.
12Recombination frequency Total amount of recombinantsƟ =Total amount of recombinants + Total amount of non-recombinantsParentGametesTheta50% non-rec and 50% rec0.5ABab90% non-rec and 10% rec0.1To quantify this distance, the recombination frequency, named Ɵ , has been defined.99% non-rec and 1% rec0.01100% non-rec
14the genetic length over which one crossover occurs in 1% of Genetic distanceGenetic distance =the genetic length over which one crossover occurs in 1% ofmeiosis. This distance is expressed in cMorgan.1 cMorgan = 0.01 recombinants = average of 1Mb (physical distance)(Assuming that the recombination frequency is uniform along the chromosomes)As double recombinants occur the further two loci are, the frequency ofrecombination does not increase proportionately.centiMorgan, named after Thomas Hunt Morgan, who first observed genetic crossing over in Drosophila. For the small distances, the amount of recombinations between two loci is indeed a measure for the genetic distance, this till 15 cM. Above 15 cM this rule is not reliable anymore, because of double crossovers
15recombination in meiosis haH96 non-rec80 non-recAbaB4 rec.15 recAH5 double rec
16Conclusion :Values of theta or genetic distance are only reliableif two loci are in the proximity of each other (max of 10 cM)
17Physical dist. Genetic dist. Chromosome 1 : 283 Mb 270 cM (0 Physical dist. Genetic dist. Chromosome 1 : 283 Mb 270 cM (0.95 cM/Mb) q arm of chromosome 21: 30 Mb 62 cM (2.1 cM/Mb) Human genome 3200 Mb 3615 cM (1.13 cM/Mb) Female genome 4460 cM Male genome 2590 cMThe genetic distance of the female chromosome is 70 percent greater than the male chromosome. Reason for this is unknown.
18Linkage equilibrium and disequilibrium Another characteristic of the genetic landscape is the phenomenon known as linkage disequilibrium.
19- Ratio form genetic distance to basepairs range from 0 - Ratio form genetic distance to basepairs range from 0.01cM/Mb to 60 cM/Mb
20Reich et al. Nature Genetics May 2001 rather large blocks of LD interspersed withrecombination hot spots
23Linkage equilibrium and disequilibrium 90% of all SNPs are shared among disparate populationsAfrican populations have smallers blocks (average 7.3kb) comparedwith 16.3kb in Europeans whereas the Chinese and Japanese blockshave an average size of 13.2kb.
25Mapping human genes by linkage analyisis Now these characteristics of the genetic landscape are elaborated, we can start with second part dedicated on mapping genes by linkage analysis. As mentioned before, my colleague Jan Hellemans will teach you how you can perform this analysis in practise by analysing genetic markers followed by an interpretation of the data and a computerized calculation. However, I wil explain you the theory behind linkage analysis and learn you how you can calculate the LOD score for simple pedigrees.
26Linkage analysis is a method that is used to decide if two loci or a loci and a disease gene are linked :Ascertain whether the recombination fraction theta between twoloci deviates significantly from 0.5.2. If theta is different from 0.5, we need to make the best estimateof theta, since this parameter tells us how close the linked loci are.Linkage is expressed as a LOD score (Z); a “ logarithm of odds”Likelihood of linkageLOD score (Ɵ) = log10Likelihood that loci are unlinked (theta = 0.5)
27Positive values of Z at a given Ɵ suggest that two loci are linked. Negative values of Z at a given Ɵ suggest that two loci are not linked.By convention, a LOD score of +3 or greater is considered definitiveevidence that two loci are linked. A LOD score below -2 excludeslinkage.Log1Probability of a recombination is ƟN is amount of recombinants in pedigreeProbability that no recombination will occur is (1-Ɵ)M is amount of non-recombinants in pedigreeƟN (1-Ɵ)MƟN (1-Ɵ)MZ (Ɵ) = log10+ log10=log10(0.5)N (0.5)M(0.5)N (0.5)M
36Interpreting LOD plots Z = log10Ɵ0 (1-Ɵ)3(0.5)0 (0.5)3=0.903 (Ɵ=0)Z = log101/2Ɵ3 + 1/2(1-Ɵ)3(0.5)3=0.602 (Ɵ=0)Strength of evidence for linkage (8 to 1) is twice as great in the phase-knownsituation compared to the phase-unknown situation.