We propose to develop a comprehensive physical map for human chromosome 3 consisting of YAC contigs and molecular probes ordered by pulsed field gel (PFG) electrophoresis. This competing renewal represents the fusion of two independent R01s, and reflects a long-standing and extremely productive corroboration between the laboratories. Our initial studies have resulted in the isolation of over 500 probes from NotI and random libraries, their localization using an extensive somatic cell hybrid mapping panel containing many of the known chromosome 3 specific rearrangements associated with malignant and developmental disorders, and the development of PFG maps for several regions of the chromosome. We have developed a rapid and economical method for screening YAC libraries which greatly facilitates the isolation of those YACs which correspond to regionally localized probes. Additional compartmentalization is provided by a set of single and two-fragment radiation reduction hybrids that will permit high resolution regional mapping to the 5-Mb level. A subset of highly polymorphic (CA)n containing probes, defining sequence-tagged sites (STS), will also be integrated into the developing physical map and contigs. Our experience with the construction of long range restriction maps using pulsed field gel electrophoresis, together with more recent experience isolating YAC clones, has led us to an important conclusion. While neither PFG maps or YAC contigs alone are sufficient to generate a comprehensive physical map necessary for completion of the first phase of the Genome Initiative, the combination of the two approaches is extremely powerful and can rapidly lead to completion. These studies will also provide the materials for the rapid isolation of disease gents encoded on this chromosome, and will facilitate our understanding of the relationship between structure and function of the human genome.
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