Sequence variability and candidate gene analysis in complex disease: association of mu opioid receptor gene variation with substance dependence

To analyze candidate genes and establish complex genotype-phenotype relatio nships against a background of high natural genome sequence variability, we have developed approaches to (i) compare candidate gene sequence informati on in multiple individuals; (ii)predict haplotypes from numerous varian ts; and (iii) classify haplotypes and identify specific sequence variants, or combinations of variants (pattern), associated with the phenotype, Using th e human mu opioid receptor gene (OPRM1) as a model system, we have combined these approaches to test a potential role of OPRM1 in substance (heroin/co caine) dependence, All known functionally relevant regions of this prime ca ndidate gene were analyzed by multiplex sequence comparison in 250 cases an d controls; 43 variants were identified and 52 different haplotypes predict ed in the subgroup of 172 African-Americans. These haplotypes were classifi ed by similarity clustering into two functionally related categories, one o f which was significantly more frequent in substance-dependent individuals, Common to this category was a characteristic pattern of sequence variants [-1793T-->A, -1699Tins, -1320A-->G, -111C-->T, +17C-->T (A6V)], which was a ssociated with substance dependence. This study provides an example of appr oaches; that have been successfully applied to the establishment of complex genotype-phenotype relationships in the presence of abundant DNA sequence variation.