COSMID ASSEMBLY AND ANCHORING TO HUMAN-CHROMOSOME-21

A human chromosome al-specific cosmid library from the Lawrence Liverm ore National Laboratory has been analyzed by two complementary methods , fingerprinting and hybridization; 40% coverage of the entire chromos ome 21 has been achieved. To prepare a contig pool, approximately 9300 cosmid clones randomly selected from the library were fingerprinted a nd automatically assembled into 467 overlapping sets by the fluorescen ce-tagged restriction fragment method. The average size of the overlap ping sets was 9.5 cosmids with minimal tiling paths consisting of 5.4 cosmids with a 10-kb extension each. However, as many as 10% of overla ps within members were estimated to be false. For regional localizatio n, we hybridized gridded arrays of cosmids with inter-ALu-PCR probes o btained from YAC clones and somatic cell hybrids and assigned 592 cosm ids to 26 subregions of 21q. Of these, 371 clones were incorporated in to 139 contigs, anchoring the total 1864 cosmids to the subregion. The remaining 221 clones were mapped as orphans. To correlate the cytogen etic, YAC, and cosmid maps on 21q, the translocation breakpoints of th e chromosomes contained in the somatic cell hybrids were mapped with r espect to the STS content of the YACs. From the gene cluster regions, 176 ribosomal and 25 alphoid clones were isolated by hybridization. To gether, these sets of anchored contigs and cosmids will provide a valu able resource for construction of a high-resolution map and for isolat ion of genes of interest from chromosome 21. (C) 1995 Academic Press, Inc.