Gene Targeting for Somatic Cell Manipulation

Abstract

Recent technological advances, not least in somatic cell genetics, have accelerated progress in human genome mapping and reverse genetics. In particular, chromosome mediated gene transfer (CMGT) (1) and irradiation and fusion gene transfer (IFGT) (2) have been exploited to great effect for enrichment cloning of predetermined regions of the human genome. Both methods allow subchromosomal fragments of human DNA (generally 1 to 50 Mbp in size) to be isolated onto a rodent cell background as a resource for DNA cloning and fine structure mapping. The success of these methods relies on a selectable marker mapping close to or within the chromosomal region of interest. A rather limited number of biochemical selection systems can be used to isolate specific regions of the genome, e.g., hypoxanthine phosphoribosyl transferase (HPRT) for Xqter and thymidine kinase (TK) for 17q. The oncogenes provide a further, novel, and widespread class of selectable marker to isolate loci involved in human cancer and linked chromosomal regions (3). In addition, antibodies directed against cell surface antigens (4) can be used to select for cells containing the corresponding chromosomal fragments. However, despite the possibilities described above, no endogenous selectable marker is known for a substantial portion of the human genome. We describe here a solution to this limitation by targeting an exogenous domi-