A 4.5-MEGABASE YEAST ARTIFICIAL CHROMOSOME CONTIG FROM HUMAN-CHROMOSOME 13Q14.3 ORDERING 9-POLYMORPHIC MICROSATELLITES (22-SEQUENCE-TAGGED SITES) TIGHTLY LINKED TO THE WILSON DISEASE LOCUS

We have previously performed a genetic analysis of multiply affected f amilies to map a locus responsible for Wilson disease (WND) to a 0.3-c entimorgan (cM) region within chromosome 13q14.3, between D13S31 and D 13S59. Here we describe the construction of a contig of almost-equal-t o 4.5 Mb, which spans this region and extends from D13S25 to D13S59. T his contig consists of 28 genomic yeast artificial chromosome (YAC) cl ones. Five critical crossover events have been defined in this interva l in two unaffected (Centre d'Etudes du Polymorphisme Humain) and thre e WND families. The combination of sequence tagged site content mappin g of YACs with both polymorphic and nonpolymorphic markers and recombi nation breakpoint mapping resulted in the following order of polymorph ic markers: 3S228-AFM238vc3-D13S133-AFM084xc5-D13S137-D13S169, D13S155 -D13S59-telomere. The recombination/physical distance ratio varies fro m almost-equal-to 3000 kb per cM in the region between D13S31 and D13S 25 to 6000 kb per cM in the region between D13S31 and D13S59. Three WN D families exhibiting recombination between the disease locus and D13S 31 or D13S59 were genotyped for additional markers in this region and further refined the location of the WND gene to between D13S155 and D1 3S133. Nine of the markers in this region of <1 cM are polymorphic mic rosatellites (seven have observed heterozygosities of 70% or above) th at will be extremely useful in prenatal and preclinical diagnosis of t his disease. This physical map is an essential step in the isolation o f the WND gene and is a framework for the identification of candidate genes.