Isolates and their potential use in complex gene mapping efforts

Linkage disequilibrium (LD), detectable with microsatellites in disease alleles over wide genetic intervals in population isolates, has facilitated mapping and positional cloning of numerous disease genes. We, among others, have shown that the LD intervals reach up to 1 Mb in general alleles of young subisolates, and that this feature most probably offers an avenue for the initial locus positioning for complex traits. Development of efficient SNP genotyping and characterization of haploblock structure of the human genome have introduced new prospects to LD-based fine mapping and haplotype-association studies. Encouraging associations have been reported for several complex diseases. Final breakthroughs in mapping of complex disease loci have emerged on large pedigrees in population isolates. Conversely, ignoring genealogical makeup of the study population seems to disclose false negative and false positive associations, directing resources down the drain.     

Mapping Genes of Complex Disease in Genetic Isolates of Daghestan

Original results of the analysis of genetic linkage between some genomic markers and two complex clinical phenotypes, schizophrenia and mental retardation, in pedigrees from Daghestan genetic isolates are described. Interpopulation differences in the epidemiology of the complex phenotypes were studied and in their genetic linkage was demonstrated. These differences are evidently related to the genetic structure of the isolates determined by their demographic history. The epidemiological index MR characterizing the lifetime morbid risk of schizophrenia varies in the Daghestan isolates studied from 0 to 4.95%, which is almost five times higher than the average worldwide population rate, 1%. Comparative genetic mapping in different isolates permitted determination of the most probable genetic linkages and associations of loci in chromosomal regions 17p11.1–12, 3q13.3, and a locus from 22q with schizophrenia and locus 12q23 with mental retardation. There is evidence that this approach is effective for detailed study of the relationship between the genetic (allele and locus) and clinical heterogeneity of complex diseases, which favors successful identification of the genes determining them. The study of linkage disequilibrium (LD) in genetic isolates of Daghestan ethnic populations (which have a common genetic background) may be an effective methodological approach for revealing the numerous contradictory results of mapping of genes of the same complex disease performed by different researchers in different regions of the world.     

Mapping genes of complex diseases in genetic isolates of Dagestan

Original results of the analysis of genetic linkage between some genomic markers and two complex clinical phenotypes, schizophrenia and mental retardation, in pedigrees from Dagestan genetic isolates are described. Interpopulation differences in the epidemiology of the complex phenotypes were studied and in their genetic linkage was demonstrated. These differences are evidently related to the genetic structure of the isolates determined by their genetic history. The MR epidemiological index characterizing the lifetime morbid risk of schizophrenia varies in the Dagestan isolates studied from 0 to 4.95%, which is almost five times higher than the average worldwide population rate, 1%. Comparative genetic mapping permitted determination of the most probable genetic linkages and associations of loci from chromosomal regions 17p11.1-12, 3q13.3, and a locus from 22q with schizophrenia and locus 12q23 with mental retardation. There is evidence that this approach is effective for detailed study of the relationship between the genetic (allele and locus) and clinical heterogeneity of complex diseases, which favors successful identification of the genes determining them. The study of linkage disequilibrium (LD) in genetic isolates of Daghestan populations (which have a common genetic background) may be an effective methodological approach for revealing the numerous contradictory results of mapping of the same genes of complex disease performed by different researchers in different regions of the world.     

Mapping genes for common diseases: the case for genetic (LD) maps

We examine the current effort to develop a haplotype map of the human genome and suggest an alternative approach which represents linkage disequilibrium patterns in the form of a metric LD map. LD maps have some of the useful properties of genetic linkage maps but have a much higher resolution which is optimal for SNP-based association mapping of common diseases. The studies that have been undertaken to date suggest that LD and recombination maps show some close similarities because of abundant, narrow, recombination hot spots. These hot spots are co-localised in all populations but, unlike linkage maps, LD maps differ in scale for different populations because of differences in population history. The prospects for developing optimized panels of SNPs and the use of linkage disequilibrium maps in disease gene localisation are assessed in the light of recent evidence.     

Short tandem-repeat polymorphism/alu haplotype variation at the PLAT locus: implications for modern human origins

Two dinucleotide short tandem-repeat polymorphisms (STRPs) and a polymorphic Alu element spanning a 22-kb region of the PLAT locus on chromosome 8p12-q11.2 were typed in 1,287-1,420 individuals originating from 30 geographically diverse human populations, as well as in 29 great apes. These data were analyzed as haplotypes consisting of each of the dinucleotide repeats and the flanking Alu insertion/deletion polymorphism. The global pattern of STRP/Alu haplotype variation and linkage disequilibrium (LD) is informative for the reconstruction of human evolutionary history. Sub-Saharan African populations have high levels of haplotype diversity within and between populations, relative to non-Africans, and have highly divergent patterns of LD. Non-African populations have both a subset of the haplotype diversity present in Africa and a distinct pattern of LD. The pattern of haplotype variation and LD observed at the PLAT locus suggests a recent common ancestry of non-African populations, from a small population originating in eastern Africa. These data indicate that, throughout much of modern human history, sub-Saharan Africa has maintained both a large effective population size and a high level of population substructure. Additionally, Papua New Guinean and Micronesian populations have rare haplotypes observed otherwise only in African populations, suggesting ancient gene flow from Africa into Papua New Guinea, as well as gene flow between Melanesian and Micronesian populations.     

Development and characterization of novel microsatellite markers from the Chinese rufous horseshoe bat (Rhinolophus sinicus) with cross-species amplification in closely related taxa

We isolated nine polymorphic microsatellite markers from the Chinese rufous horseshoe bat (Rhinolophus sinicus) using an enriched library method. We assessed genetic polymorphism at these loci in 42 individuals from a single population. We recorded high genetic diversity with four to 17 alleles per locus, and estimated expected and observed heterozygosity values ranging from 0.492 to 0.910 and from 0.462 to 0.881, respectively. No locus departed from Hardy-Weinberg equilibrium following Bonferroni correction, and no linkage disequilibrium was detected. Most loci successfully cross-amplified congeneric species. These loci will be used to characterize phylogeographical history of Rhinolophus sinicus in China.     

Extent and distribution of linkage disequilibrium in three genomic regions

The positional cloning of genes underlying common complex diseases relies on the identification of linkage disequilibrium (LD) between genetic markers and disease. We have examined 127 polymorphisms in three genomic regions in a sample of 575 chromosomes from unrelated individuals of British ancestry. To establish phase, 800 individuals were genotyped in 160 families. The fine structure of LD was found to be highly irregular. Forty-five percent of the variation in disequilibrium measures could be explained by physical distance. Additional factors, such as allele frequency, type of polymorphism, and genomic location, explained <5% of the variation. Nevertheless, disequilibrium was occasionally detectable at 500 kb and was present for over one-half of marker pairs separated by <50 kb. Although these findings are encouraging for the prospects of a genomewide LD map, they suggest caution in interpreting localization due to allelic association.     

Genetic isolates in East Asia: a study of linkage disequilibrium in the X chromosome

The background linkage disequilibrium (LD) in genetic isolates is of great interest in human genetics. Although many empirical studies have evaluated the background LD in European isolates, such as the Finnish and Sardinians, few data from other regions, such as Asia, have been reported. To evaluate the extent of background LD in East Asian genetic isolates, we analyzed the X chromosome in the Japanese population and in four Mongolian populations (Khalkh, Khoton, Uriankhai, and Zakhchin), the demographic histories of which are quite different from one another. Fisher's exact test revealed that the Japanese and Khalkh, which are the expanded populations, had the same or a relatively higher level of LD than did the Finnish, European American, and Sardinian populations. In contrast, the Khoton, Uriankhai, and Zakhchin populations, which have kept their population size constant, had a higher background LD. These results were consistent with previous genetic anthropological studies in European isolates and indicate that the Japanese and Khalkh populations could be utilized in the fine mapping of both complex and monogenic diseases, whereas the Khoton, Uriankhai, and Zakhchin populations could play an important role in the initial mapping of complex disease genes.     

Recombination disequilibrium in ideal and natural populations

Following Hardy-Weinberg disequilibrium (HWD) occurring at a single locus and linkage disequilibrium (LD) between two loci in generations, we here proposed the third genetic disequilibrium in a population: recombination disequilibrium (RD). RD is a measurement of crossover interference among multiple loci in a random mating population. In natural populations besides recombination interference, RD may also be due to selection, mutation, gene conversion, drift and/or migration. Therefore, similarly to LD, RD will also reflect the history of natural selection and mutation. In breeding populations, RD purely results from recombination interference and hence can be used to build or evaluate and correct a linkage map. Practical examples from F₂, testcross and human populations indeed demonstrate that RD is useful for measuring recombination interference between two short intervals and evaluating linkage maps. As with LD, RD will be important for studying genetic mapping, association of haplotypes with disease, plant breading and population history.     

Haplotypes at ATM identify coding-sequence variation and indicate a region of extensive linkage disequilibrium

Genetic variation in the human population may lead to functional variants of genes that contribute to risk for common chronic diseases such as cancer. In an effort to detect such possible predisposing variants, we constructed haplotypes for a candidate gene and tested their efficacy in association studies. We developed haplotypes consisting of 14 biallelic neutral-sequence variants that span 142 kb of the ATM locus. ATM is the gene responsible for the autosomal recessive disease ataxia-telangiectasia (AT). These ATM noncoding single-nucleotide polymorphisms (SNPs) were genotyped in nine CEPH families (89 individuals) and in 260 DNA samples from four different ethnic origins. Analysis of these data with an expectation-maximization algorithm revealed 22 haplotypes at this locus, with three major haplotypes having frequencies > or = .10. Tests for recombination and linkage disequilibrium (LD) show reduced recombination and extensive LD at the ATM locus, in all four ethnic groups studied. The most striking example was found in the study population of European ancestry, in which no evidence for recombination could be discerned. The potential of ATM haplotypes for detection of genetic variants through association studies was tested by analysis of 84 individuals carrying one of three ATM coding SNPs. Each coding SNP was detected by association with an ATM haplotype. We demonstrate that association studies with haplotypes for candidate genes have significant potential for the detection of genetic backgrounds that contribute to disease.     

Use of population isolates for mapping complex traits

Geneticists have repeatedly turned to population isolates for mapping and cloning Mendelian disease genes. Population isolates possess many advantages in this regard. Foremost among these is the tendency for affected individuals to share ancestral haplotypes derived from a handful of founders. These haplotype signatures have guided scientists in the fine mapping of scores of rare disease genes. The past successes with Mendelian disorders using population isolates have prompted unprecedented interest among medical researchers in both the public and private sectors. Despite the obvious genetic and environmental complications, geneticists have targeted several population isolates for mapping genes for complex diseases.     

Assignment of the locus for a new lethal neonatal metabolic syndrome to 2q33-37.

A new neonatal syndrome characterized by intrauterine growth retardation, lactic acidosis, aminoaciduria, liver hemosiderosis, and early death was recently described. The pathogenesis of this disease is unknown. The mode of inheritance is autosomal recessive, and so far only 17 cases have been reported in 12 Finnish families. Here we report the assignment of the locus for this new disease to a restricted region on chromosome 2q33-37. We mapped the disease locus in a family material insufficient for traditional linkage analysis by using linkage disequilibrium, a possibility available in genetic isolates such as Finland. The primary screening of the genome was performed with samples from nine affected individuals in five families. In the next step, conventional linkage analysis was performed in eight families, with a total of 12 affected infants, and finally the locus assignment was proved by demonstrating linkage disequilibrium to the regional markers in 20 disease chromosomes. Linkage analysis restricted the disease locus to a 3-cM region between markers D2S164 and D2S2359, and linkage disequilibrium with the ancestral haplotype restricted the disease locus further to the immediate vicinity of marker D2S2250.     

Genome-wide association filtering using a highly locus-specific transmission/disequilibrium test

Multimarker transmission/disequilibrium tests (TDTs) are powerful association and linkage tests used to perform genome-wide filtering in the search for disease susceptibility loci. In contrast to case/control studies, they have a low rate of false positives for population stratification and admixture. However, the length of a region found in association with a disease is usually very large because of linkage disequilibrium (LD). Here, we define a multimarker proportional TDT (mTDT _P ) designed to improve locus specificity in complex diseases that has good power compared to the most powerful multimarker TDTs. The test is a simple generalization of a multimarker TDT in which haplotype frequencies are used to weight the effect that each haplotype has on the whole measure. Two concepts underlie the features of the metric: the ‘common disease, common variant’ hypothesis and the decrease in LD with chromosomal distance. Because of this decrease, the frequency of haplotypes in strong LD with common disease variants decreases with increasing distance from the disease susceptibility locus. Thus, our haplotype proportional test has higher locus specificity than common multimarker TDTs that assume a uniform distribution of haplotype probabilities. Because of the common variant hypothesis, risk haplotypes at a given locus are relatively frequent and a metric that weights partial results for each haplotype by its frequency will be as powerful as the most powerful multimarker TDTs. Simulations and real data sets demonstrate that the test has good power compared with the best tests but has remarkably higher locus specificity, so that the association rate decreases at a higher rate with distance from a disease susceptibility or disease protective locus.     

Joint modeling of linkage and association: identifying SNPs responsible for a linkage signal

Once genetic linkage has been identified for a complex disease, the next step is often association analysis, in which single-nucleotide polymorphisms (SNPs) within the linkage region are genotyped and tested for association with the disease. If a SNP shows evidence of association, it is useful to know whether the linkage result can be explained, in part or in full, by the candidate SNP. We propose a novel approach that quantifies the degree of linkage disequilibrium (LD) between the candidate SNP and the putative disease locus through joint modeling of linkage and association. We describe a simple likelihood of the marker data conditional on the trait data for a sample of affected sib pairs, with disease penetrances and disease-SNP haplotype frequencies as parameters. We estimate model parameters by maximum likelihood and propose two likelihood-ratio tests to characterize the relationship of the candidate SNP and the disease locus. The first test assesses whether the candidate SNP and the disease locus are in linkage equilibrium so that the SNP plays no causal role in the linkage signal. The second test assesses whether the candidate SNP and the disease locus are in complete LD so that the SNP or a marker in complete LD with it may account fully for the linkage signal. Our method also yields a genetic model that includes parameter estimates for disease-SNP haplotype frequencies and the degree of disease-SNP LD. Our method provides a new tool for detecting linkage and association and can be extended to study designs that include unaffected family members.     

Design and sample-size considerations in the detection of linkage disequilibrium with a disease locus.

The presence of linkage disequilibrium between closely linked loci can aid in the fine mapping of disease loci. We investigate the power of several designs for sampling individuals with different disease genotypes. As expected, haplotype data provide the greatest power for detecting disequilibrium, but, in the absence of parental information to resolve the phase of double heterozygotes, the most powerful design samples only individuals homozygous at the trait locus. For rare diseases, such a scheme is generally not feasible, and we also provide power and sample-size calculations for designs that sample heterozygotes. The results provide information useful in planning disequilibrium studies.     

Gene mapping by linkage and association analysis

Genetic analysis is used to map genes, including disease loci, to positions within the human genome. Linkage analysis depends on the co-segregation of a gene (locus) and a phenotype through a pedigree, while association analysis, or linkage disequilibrium mapping, depends on measuring deviation from the random occurrence of alleles in a haplotype in unrelated individuals or nuclear families. Complex computer programs may be used in both forms of analysis. In recent years most interest has focused on identifying genes involved in common, multifactorial diseases. Here I review some current and developing techniques of genetic analysis and give references to where further information can be obtained.     

Hierarchical modeling of linkage disequilibrium: genetic structure and spatial relations

Linkage disequilibrium (LD) mapping offers much promise for the positional cloning of disease-causing genes. However, conventional estimates of LD may fluctuate substantially across contiguous genomic regions, because of population-specific phenomena such as mutation, genetic drift, population structure, and variations in allele frequencies. This fluctuation makes it difficult to interpret patterns of LD and distinguish where a causal gene is located. To address this issue, we propose hierarchical modeling of LD (HLD) for fine-scale mapping. This approach incorporates information on haplotype block structure and chromosomal spatial relations to refine the pattern of LD, increasing the ability to localize disease genes. Here, we present a framework for HLD, a simulation study assessing the performance of HLD under various scenarios, and an application of HLD to existing data. This work demonstrates that hierarchical modeling of linkage disequilibrium is a valuable and flexible approach for fine-scale mapping.     

A coalescent approach to study linkage disequilibrium between single-nucleotide polymorphisms

We present the results of extensive simulations that emulate the development and distribution of linkage disequilibrium (LD) between single-nucleotide polymorphisms (SNPs) and a gene locus that is phenotypically stratified into two classes (disease phenotype and wild-type phenotype). Our approach, based on coalescence theory, allows an explicit modeling of the demographic history of the population without conditioning on the age of the mutation, and serves as an efficient tool to carry out simulations. More specifically, we compare the influence that a constant population size or an exponentially growing population has on the amount of LD. These results indicate that attempts to locate single disease genes are most likely successful in small and constant populations. On the other hand, if we consider an exponentially growing population that started to expand from an initially constant population of reasonable size, then our simulations indicate a lower success rate. The power to detect association is enhanced if haplotypes constructed from several SNPs are used as markers. The versatility of the coalescence approach also allows the analysis of other relevant factors that influence the chances that a disease gene will be located. We show that several alleles leading to the same disease have no substantial influence on the amount of LD, as long as the differences between the disease-causing alleles are confined to the same region of the gene locus and as long as each allele occurs in an appreciable frequency. Our simulations indicate that mapping of less-frequent diseases is more likely to be successful. Moreover, we show that successful attempts to map complex diseases depend crucially on the phenotype-genotype correlations of all alleles at the disease locus. An analysis of lipoprotein lipase data indicates that our simulations capture the major features of LD occurring in biological data.     

Populationsgenetik des humanen X-Chromosoms

Mitochondrial DNA and the Y chromosome (ChrY) are both highly informative regarding human evolution, demographic history, and the genetic relationships between extant populations. The major reason for this is that both genomic compartments do not recombine, except for the pseudo-autosomal regions of ChrY, and that typing of haploid markers automatically allows the identification of haplotypes. In terms of its recombination behaviour, the X chromosome (ChrX) falls between autosomes and ChrY. The significance of ChrX in terms of population genetics is partially based on the fact that its haplotypes are easier to determine than those of autosomes. While ChrY and mtDNA each represent a single locus only, with a common evolutionary history of all their constituents, ChrX comprises several regions that may each reflect its own history. Therefore, ChrX studies seem most suitable for distinguishing between subpopulations or for research on regional ethnic structures. The analysis of linkage disequilibrium (LD) is one of the key aspects of population genetics studies of ChrX markers because LD may be an indicator of genetic isolation or of the emergence from a small founder population. Populations with high LD play an important role in the identification of genes involved in the aetiology of multifactorial diseases.     

Genome screens using linkage disequilibrium tests: optimal marker characteristics and feasibility.

Linkage disequilibrium (LD) testing has become a popular and effective method of fine-scale disease-gene localization. It has been proposed that LD testing could also be used for genome screening, particularly as dense maps of diallelic markers become available and automation allows inexpensive genotyping of diallelic markers. We compare diallelic markers and multiallelic markers in terms of sample sizes required for detection of LD, by use of a single marker locus in a case-control study, for rare monophyletic diseases with Mendelian inheritance. We extrapolate from our results to discuss the feasibility of single-marker LD screening in more-complex situations. We have used a deterministic population genetic model to calculate the expected power to detect LD as a function of marker density, age of mutation, number of marker alleles, mode of inheritance of a rare disease, and sample size. Our calculations show that multiallelic markers always have more power to detect LD than do diallelic markers (under otherwise equivalent conditions) and that the ratio of the number of diallelic to the number of multiallelic markers needed for equivalent power increases with mutation age and complexity of mode of inheritance. Power equivalent to that achieved by a multiallelic screen can theoretically be achieved by use of a more dense diallelic screen, but mapping panels of the necessary resolution are not currently available and may be difficult to achieve. Genome screening that uses single-marker LD testing may therefore be feasible only for young (<20 generations), rare, monophyletic Mendelian diseases, such as may be found in rapidly growing genetic isolates.