Ordering of Chromosome Specific Overlapping Cosmid Clones

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In the phase II work proposed research we will determine the overlapping relationships between-10,000 clones from a cosmid library derived from partially-digested, chromosome-sorted human DNA. This information will allow the creation of an ordered chromosome specific library. This library, and others to follow, will be indispensable tools for the human genome mapping and sequencing project. The phase II work will create an ordered cosmid library as well as kits, devices, and instruction manuals that can be used by other investigators to accomplish the same result on segments of DNA as large as entire human chromosomes. The phase II work involves cross-hybridization of the inserts from a large number of cosmid clones to each other independently in all combinations using the "pen-line" method developed in phase I work. The "pen-line" method allows discrete hybridization of all cloned DNAs separately and independently to all probes generated from the same set clones and, therefore, allows for simultaneous determination of the overlap between all clones in the set. The method is several fold faster than current repeatable protocol. In phase II work, we propose to use the method on an overlapping clone library derived from chromosome 20.