Thymidylate synthase-deficient Chinese hamster cells: a selection system for human chromosome 18 and experimental system for the study of thymidylate synthase regulation and fragile X expression.
Chinese hamster lung (CHL) V79 cells already deficient in hypoxanthine phosphoribosyltransferase were exposed to uv light and selected for mutations causing deficiency of thymidylate synthase ( TS) by their resistance to aminopterin in the presence of thymidine and limiting amounts of methyl tetrahydrofolate. Three of seven colonies chosen for initial study were shown to be thymidylate synthase deficient ( TS-) by enzyme assay, thymidine auxotrophy, and their inability to incorporate labeled deoxyuridine into their DNA in vivo. Complementation analysis of human X TS- hamster hybrids revealed that TS activity segregated with human chromosome 18. Southern analysis of a panel of 14 human X hamster hybrids probed with complementary DNA from mouse TS confirmed the chromosome assignment of TS to human chromosome 18; quantitative Southern blotting using unbalanced human cell lines further localized the gene to 18q21.31----qter. Another hybrid was generated that contained a human X chromosome with the Xq28 folate-dependent fragile site as its only human chromosome in a hamster TS- background. The fragile site could be easily and reproducibly expressed in this hybrid without the use of antimetabolites simply by removing exogenous thymidine from the medium. These TS-deficient cells are useful for: somatic cell genetics as a unique selectable marker for human chromosome 18, studies on regulation of the TS gene, and analysis of the fragile (X) chromosome and other folate-dependent fragile sites.[1]References
- Thymidylate synthase-deficient Chinese hamster cells: a selection system for human chromosome 18 and experimental system for the study of thymidylate synthase regulation and fragile X expression. Nussbaum, R.L., Walmsley, R.M., Lesko, J.G., Airhart, S.D., Ledbetter, D.H. Am. J. Hum. Genet. (1985) [Pubmed]
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