CONSTRUCTION OF A YEAST ARTIFICIAL CHROMOSOME CONTIG SPANNING THE SPINAL MUSCULAR-ATROPHY DISEASE GENE REGION

The childhood spinal muscular atrophies (SMAs) are the most common, se rious neuromuscular disorders of childhood second to Duchenne muscular dystrophy. A single locus for these disorders has been mapped by reco mbination events to a region of 0.7 centimorgan (range, 0.1-2.1 centim organs) between loci D5S435 and MAP1B on chromosome 5q11.2-13.3. By us ing PCR amplification to screen yeast artificial chromosome (YAC) DNA pools and the PCR-vectorette method to amplify YAC ends, a YAC contig was constructed across the disease gene region. Nine walk steps identi fied 32 YACs, including a minimum of seven overlapping YAC clones (ave rage size, 460 kb) that span the SMA region. The contig is characteriz ed by a collection of 30 YAC-end sequence tag sites together with seve n genetic markers. The entire YAC contig spans a minimum of 3.2 Mb; th e SMA locus is confined to roughly half of this region. Microsatellite markers generated along the YAC contig segregate with the SMA locus i n all families where the flanking markers (D5S435 and MAP1B) recombine . Construction of a YAC contig across the disease gene region is an es sential step in isolation of the SMA-encoding gene.