Generation of region- and species-specific expressed gene probes from somatic cell hybrids.
Human genes expressed in interspecific somatic cell hybrids can be cloned specifically by subtractive cDNA hybridization. This approach is based on the observation that cDNA fragments from noncoding segments of mature human transcripts do not form stable heteroduplexes with their rodent homologues under high-stringency hybridization conditions. Thus, small, oligo-dT primed cDNAs from a rat/human hybrid retaining a fragment of human chromosome 17 were enriched for human sequences by hybridization with RNA from a sister clone containing a smaller human chromosome fragment. The enriched probe was used to screen a human cDNA library, and nine expressed genes from within the non-overlap region were obtained. This method should be useful for cloning active human genes from defined chromosome segments.
Jones
K W
KW
Department of Molecular Medicine, Fred Hutchinson Cancer Research Center, Seattle, Washington 98104.
Chevrette
M
M
Shapero
M H
MH
Fournier
R E
RE
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002478:Cells, Cultured; D002871:Chromosome Banding; D002874:Chromosome Mapping / Q000379:methods; D002886:Chromosomes, Human, Pair 17; D015342:DNA Probes; D005347:Fibroblasts / Q000166:cytology / Q000502:physiology; D015723:Gene Library; D008040:Genetic Linkage; D006801:Humans; D006822:Hybrid Cells / Q000502:physiology*; D008114:Liver Neoplasms, Experimental; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D051381:Rats; D012689:Sequence Homology, Nucleic Acid; D014404:Tubulin / Q000235:genetics; D014407:Tumor Cells, Cultured
D015342:DNA Probes; D009838:Oligodeoxyribonucleotides; D014404:Tubulin