Genomic screening to map disease loci by association requires automation, pooling of DNA samples, and 3,000-6,000 highly polymorphic, evenly spaced microsatellite markers. Case-control samples can be used in an initial screen, followed by family-based data to confirm marker associations. Association mapping is relevant to genetic studies of complex diseases in which linkage analysis may be less effective and to cases in which multigenerational data are difficult to obtain, including rare or late-onset conditions and infectious diseases. The method can also be used effectively to follow up and confirm regions identified in linkage studies or to investigate candidate disease loci. Study designs can incorporate disease heterogeneity and interaction effects by appropriate subdivision of samples before screening. Here we report use of pooled DNA amplifications-the accurate determination of marker-disease associations for both case-control and nuclear family-based data-including application of correction methods for stutter artifact and preferential amplification. These issues, combined with a discussion of both statistical power and experimental design to define the necessary requirements for detecting of disease loci while virtually eliminating false positives, suggest the feasibility and efficiency of association mapping using pooled DNA screening.

Congenital generalized lipodystrophy (CGL, Berardinelli-Seip Syndrome, OMIM # 269700) is a rare autosomal recessive disorder characterized by near complete absence of adipose tissue from birth. Affected individuals have marked insulin resistance, hypertriglyceridemia and acanthosis nigricans, and develop diabetes mellitus during teenage years. The genetic defect for CGL is unknown. A semi-automated genome-wide scan with a set of highly polymorphic short tandem repeats (STR) was carried out in 17 well-characterized pedigrees and identified a locus for CGL to chromosome 9q34. The maximum two-point lod score obtained was 3.6 at D9S1818 (theta(max) = 0.05). There was evidence for genetic heterogeneity (alpha = 0.73) and 2 of the pedigrees were unlinked. Multipoint linkage analysis excluding the 2 unlinked families yielded a peak lod score of 5.4 between loci D9S1818 and D9S1826. The CGL1 critical region harbors a plausible candidate gene encoding the retinoid X receptor alpha (RXRA) that plays a central role in adipocyte differentiation. Identification of the CGL gene(s) will contribute to our understanding of the adipocyte differentiation and elucidation of the mechanisms of insulin resistance in disorders of adipose tissue.

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