VERTICALLY INTEGRATED YAC BASED X CHROMOSOME ANALYSIS

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Vertically Integrated YAC-Based X Chromosome Analysis: In the first phase of the human genome initiative, the Human Genome Center here provided the ideas, methods, and data to achieve first-generation maps of chromosomes 7 and X. The maps are based on overlapping yeast artificial chromosomes (YACs), and are formatted with sequence-tagged sites (STSs) that are detected by the polymerase chain reaction (PCR). The Center's next goal is to achieve an extended, vertically integrated analysis that proceeds smoothly all the way to sequence. As a first step, the X chromosome map will be refined with STS markers at least every 100 kb, and will be annotated with genes and markers that detect polymorphism (Project 1). This will provide a platform for the needs of researchers seeking disease genes or studying structural or developmental biology, and will also serve as a starting framework to merge mapping with high-throughput sequencing. Data will be organized and integrated with data from outside sources in an Information Handling and Analysis Core, which will submit rationalized maps and the underlying data to international databases in a timely fashion. Materials, including clone libraries and STS primer pairs, will be stored and maintained in a Mapping Resources and Implementation Core, which will also aid in the adaptation for general use of methods and protocols coming from Project laboratories and outside sources. New technology required for the finer mapping and sequencing will be developed, including, in Project 1, the institution of more stable new YACs; development and adaptation of methods to find genes, including new types of recombination trapping of cDNAs encoded by YACs; use of degenerate STSs to find the locations of members of gene families; and adaptation of PCR-based screening to find full-length cDNAs starting from short sequences; in Project 2, testing an ordered shotgun approach to sequencing compared to the best available alternatives; and In addition, comparative genome analysis would be aided by the sequencing of selected regions in mouse and human DNA that are syntenically equivalent, and by the provision of conserved STSs to research groups studying the evolution of chromosome structure and content.