The goal of Project 2 is to convert the contig-based sequence-ready maps produced by Project 1 into detailed high-resolution annotated sequence-based maps by selected DNA sequencing. Operating in parallel with Project 1, using the set of cosmids maps generated for human chromosomes 11, 21, 20, 10, 13, 14 and 18 and 8, this project will be carried out in a highly automated manner. The sequence-ready contig maps produced by Project 1, consisting of 10 to 20X coverage of the cosmid ends, will be subjected to high-throughput DNA sequencing directly from the cosmid templates, using automated template DNA preparation, automated reaction assembly and incubation, automated sequence determination using conventional fluorescence technology, and immediate on-line sequence analysis using custom designed parallel processing microcomputers. This analysis will result in the one-pass sequence of about one-third of each chromosome in ordered sequence fragments of 350 to 550 bp in length, with gaps of 1 to 5 kb, and will serve as an immediate prelude to complete DNA sequencing. This will be the determination and analysis of 30 to 50 percent of the entire chromosome sequence. The analysis of this information using available programs and additional software to be developed by the computation core should result in: (1) The construction of STSs based upon physical position with an even spacing of at least 100 kb; (2) The detection of sufficient simple sequence repeats, and their conversion to PCR- detectable polymorphic markers, to refine the genetic map to at least 0.1 centimorgan resolution based upon physical position; and (3) The complete genomic or cDNA sequence of putative genes of exceptional interest, such as those encoding unique proteins or likely to represent candidate genes for human genetic diseases.
