High density physical mapping of chromosome 3p by hybridization of somatic cell hybrid derived Alu-PCR products
- M.I.T., Cambridge, MA (United States); and others
We have produced high density physical maps covering most of chromosome 3 using a hybridization-based approach to YAC isolation and contig assembly. The strategy has been to use a well characterized panel of somatic cell hybrids to generate probe sets distributed across the chromosome. From each of 50 somatic cell hybrids, a library of {approximately}100-600 cloned Alu-PCR products was isolated and Alu-PCR inserts from the full set of clones was spotted at high density on nylon membranes. Representative sets of unique clones from most of the hybrids were used to screen {approximately}16 genome equivalents of CEPH YACs by hybridization. These results, combined with our previous results and data from CEPH, produced contigs covering most of the chromosomes. The use of somatic cell hybrid-derived probe sets allowed easy integration of contigs with physical mapping boundaries defined by the somatic cell hybrids. We will describe various Alu-PCR strategies subsequently used to achieve closure, including successful identification of Alu-PCR products from the spotted set which lie in regions not previously covered by YAC contigs and use of Alu-PCR walking strategies to fill gaps and confirm tentative linkages. Our basic YAC screening methodology allows one individual to screen {approximately}16 genome equivalents of CEPH YACs with 96 probes/week at a material cost of less than $1 per locus. We have now mapped >2,200 loci on chromosome 3, with average interlocus distances of {approximately}50-100 kb over large regions. Alu-PCR-defined loci spaced along the chromosome at regular intervals are being converted to STSs. Our results indicate that use of a hybridization-based approach to physical mapping constitutes an efficient, accurate, high throughput method for isolating YACs and assembling YAC contigs.
- OSTI ID:
- 134473
- Report Number(s):
- CONF-941009--
- Journal Information:
- American Journal of Human Genetics, Journal Name: American Journal of Human Genetics Journal Issue: Suppl.3 Vol. 55; ISSN AJHGAG; ISSN 0002-9297
- Country of Publication:
- United States
- Language:
- English
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