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© 1994 Oxford University Press

OTHER

A YAC-based binning strategy facilitating the rapid assembly of cosmid contigs: 1.6 Mb of overlapping cosmids in Xp22

Martin C. Wapenaar1, M.Vittoria Schiaffino1, Maria T. Bassi1, Laura Schaefer1, A.Craig Chinault1,2, Huda Y. Zoghbi1,2,3 and Andrea Ballabio1,2,*

1Department of Molecular and Human Genetics, Baylor College of Medicine One Baylor Plaza, Houston, TX 77030, USA 2Human Genome Center, Baylor College of Medicine One Baylor Plaza, Houston, TX 77030, USA 3Department of Pediatrics, Baylor College of Medicine One Baylor Plaza, Houston, TX 77030, USA

*To whom correspondence should be addressed

Received April 6, 1994; Revised May 13, 1994; Accepted May 13, 1994

We have applied a yeast artificial chromosome (YAC)-based cosmid isolation and binning strategy to convert a YAC contig in Xp22 into 1.6 Mb of overlapping cosmids. This strategy is based on the screening of a high-density arrayed X chromosome-specific cosmid library with large YAC-derived restriction fragments and entire YAC probes. Cosmids selected in this way were gridded on dot blots and further mapped into bins defined by the overlap intervals of the YACs and YAC fragments. This rapid binning of cosmids simplified the subsequent assembly of cosmid contigs by restriction fingerprint hybridization. In total, we identified 139 cosmids spanning the entire 1.6 Mb region with a minimal overlap set of 53 clones. These cosmids were assigned to 17 bins and 9 contigs. One of the contigs is 665 kb in length and is one of the largest uninterrupted cosmid contigs in humans reported to date. The gaps between the contigs are minor and, together, they represent less than 7% of the region covered. Two previously identified genes are contained in these cosmids, the gene for amelogenin (AMG) and the recently isolated putative chloride channel gene CICN4. In addition, two disease loci have been mapped to this region: X-linked ocular albinism type 1 (OA1) and the microphthalmia with linear skin defects (MLS) syndrome. The assembly of the cosmid maps allowed us to determine the size of the deletion intervals for these two loci, which were estimated to be 110 kb for OA1 and 570 kb for MLS. These cosmid contigs will greatly facilitate the positional cloning of the OA1 and MLS disease genes. Together with the Huntington disease gene region on chromosome 4, this region in Xp22 represents one of the best characterized large regions in the human genome.


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