Construction of human chromosome-3-specific radiation hybrids and characterization by Alu-PCR

Using a modification of the procedure developed by Cox et al. [Genomics 4 (1989) 397-407], we isolated and characterized 60 radiation hybrids (RH) prepared by fusing an X-ray-irradiated Chinese hamster-human chromosome 3 (Chr 3) cell line (Q314-2) with a UrdA Chinese hamster mutant cell line. The RH were screened for human DNA content by PCR amplification using primers directed to the human Alu repeat sequences. Over 80% (50/60) were scored as positive for the retention of human DNA. Of them, 18 were characterized with Chr-3-specific single-copy DNA probes of known map location. These experiments demonstrated that the RH analyzed contained distinct subregions of human Chr 3. The RH that we have produced constitute a bank of cellular clones containing small segments of Chr 3. In the accompanying paper [Atchison et al., Gene 151 (1994) 325-328], we present the construction of rare-restriction-site linking libraries and the sequence tagged site characterization of in situ localized clones.