Summary
Mutants resistant to 6-thioguanine were selected from CHO cells which were either temperature sensitive or proline requiring. These mutants were stable and had low levels of hypoxanthine guanine phosphoribosyl transferase (HGPRT). Hybrids were selected which were heteroallelic at the hgprt locus and complementation between the mutants used was not observed. Interallelic recombination at this locus would generate hgprt + cells which could be selected in Littlefield's HAT medium. Selection experiments with hybrids containing three different pairs of mutants yielded no recombinants among populations of 4x106-2x107 cells. After treatment with the recombinagen mitomycin C, 3 putative recombinants were detected amongst 1.4x107 surviving cells from one hybrid. One of these strains was examined and shown to have a normal level of HGPRT and its heterozygosity at this locus was demonstrated by the segregation of colonies resistant to 6-thioguanine. It cannot be excluded that the rare hgprt + colonies seen arose by mutation rather than by recombination. Mitotic allelic recombination therefore appears to be a much less frequent event in CHO cells than it is in lower eukaryotes. It is possible that mitotic recombination is effectively suppressed in mammalian cells to prevent the expression of deleterious recessive mutants.
Similar content being viewed by others
References
Arlett, C.F., Turnbull, D., Harcourt, S.A., Lehmann, A.R., Colella, C.M.: A comparison of the 8-azaguanine and ouabain resistance systems for the selection of induced mutant Chinese hamster cells. Mutation Res. 33, 261–278 (1975)
Beaudet, A.L., Roufa, D.J., Caskey, C.T.: Mutations affecting the structure of hypoxanthine guanine phosphoribosyltransferase in cultured Chinese hamster cells. Proc. nat. Acad. Sci. (Wash) 70 320–324 (1973)
Brown, S.M., Ritchie, D.A., Subak-Sharpe, J.H.: Genetic studies with herpes simplex virus type 1: The isolation of temperaturesensitive mutants, their arrangement into complementation groups and recombination analysis leading to a linkage map. J. gen. Virol. 18, 329–346 (1973)
Chaganti, R.S.K., Schonberg, S., German, J.: A many fold increase in sister chromatid exchanges in Bloom's syndrome lymphocytes. Proc. nat. Acad. Sci. (Wash.) 71, 4508–4512 (1974)
Chasin, L.A.: Non-linkage of induced mutations in Chinese hamster cells. Nature (Lond.) New Biol. 240, 50–52 (1972)
Chasin, L.A., Urlaub, G.: Mutant alleles for hypoxanthine phosphoribosyl transferase: codominant expression, complementation, and segregation in hybrid Chinese hamster cells. Somatic Cell Genetics 2, 453–467 (1976)
Chu, E.H.Y., Malling, H.V.: Mammalian cell genetics 11. Chemical induction of specific locus mutations in Chinese hamster cells in vitro. Proc. nat. Acad. Sci. (Wash.) 61, 1306–1312 (1968)
Cox, R.P., Kauss, M.R., Balis, M.E., Dancis, J.: Evidence for transfer of enzyme product as the basis for metabolic co-operation between tissue culture fibroblasts of Lesch-Nyhan disease and normal cells. Proc. nat. Acad. Sci. (Wash.) 67, 1573–1579 (1970)
Deaven, L.L., Petersen, D.F.: The chromosomes of CHO, an aneuploid Chinese hamster cell line: G-band, C-band and autoradiographic analyses. Chromosoma (Berl.) 41, 129–144 (1973)
DeMars, R.: Resistance of cultured human fibroblasts and other cells to purine and pyrimidine analogues in relation to mutagenesis detection. Mutation Res. 24, 335–364 (1974)
Esposito, M.S.: X-ray and meiotic fine structure mapping of the adenine 8 locus in Saccharomyces cerevisiae. Genetics 58, 507–527 (1968)
Fenwick, R.G., Caskey, C.T.: Mutant Chinese hamster cells with a thermosensitive hypoxanthine-guanine phosphoribosyl transferase. Cell 5, 115–122 (1975)
Fox, M., Boyle, J.M., Fox, B.W.: Biological and chemical characterisation of purine analogue resistant clones of V79 Chinese hamster cells. Mutation Res. 35, 289–310 (1976)
Gillin, F.D., Roufa, D.J., Beaudet, A.L., Caskey, C.T.: 8-azaguanine resistance in mammalian cells 1. Hypoxanthine-guanine phosphoribosyltransferase. Genetics 72, 239–252 (1972)
Gladstone, P., Sabo, K., Pious, L., Pious, D.: Correlation between production of quadriaradial chromosome configurations and mitotic recombinants in the HLA region in cultured lymphoid cells. Proc. V Int. Congr. Human Genetics, Excerpta medica (Amsterdam) Int. Congr. Series No. 397, p. 127. (1976)
Holliday, R.: The induction of mitotic recombination by mitomycin C in Ustilago and Saccharomyces. Genetics 50, 323–335 (1964)
Holliday, R.: Studies on mitotic gene conversion in Ustilago. Genet. Res. 8, 323–337 (1966)
Holliday, R.: Biochemical measure of the time and frequency of radiation-induced allelic recombination in Ustilago. Nature (Lond.) New Biol. 232, 233–236 (1971)
Holliday, R.: Recombination and meiosis. In: The meiotic process (ed. R. Riley, r. Flavell). Phil. Trans. (in press, 1976)
Hughes, S.H., Wahl, G.M., Capecchi, M.R.: Purification and characterization of mouse hypoxanthine guanine phosphoribosyl transferase. J. biol. Chem. 250, 120–126 (1975)
Huttner, K.M., Ruddle, F.H.: Study of mitomycin-C induced chromosomal exchange. Chromosoma (Berl.) 56, 1–13 (1976)
Klebe, R.J., Chen, T., Ruddle, F.H.: Controlled production of proliferating somatic cell hybrids. J. Cell Biol. 45, 74–81 (1970)
Latt, S.A.: Sister chromatid exchanges indices of human chromosome damage and repair: detection by fluorescence and induction by mitomycin C. Proc. nat. Acad. Sci. (Wash.) 71, 3162–3166 (1974)
Littlefield, J.W.: The use of drug resistant markers to study the hybridization of mouse fibroblasts. Exp. Cell Res. 41, 190–196 (1966)
Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J.: Protein measurement with the folin phenol reagent. J. biol. Chem. 193, 265–275 (1951)
Moorhead, P.S., Nowell, P.C., Mellman, W.J., Battips, D.M., Hungerford, D.A.: Chromosome preparations of leukocytes cultured from human peripheral blood. Exp. Cell Res. 20, 613–616 (1960)
Parag, Y., Parag, G.: Mutations affecting mitotic recombination frequency in haploids and diploids of the filamentous fungus Aspergillus nidulans Molec. gen. Genet. 137, 109–123 (1975)
Peto, R., Roe, F.J.C., Lee, P.N., Levy, L., Clack, J.: Cancer and ageing in mice and men. Brit. J. Cancer. 32, 411–426 (1975)
Pontecorvo, G.: Tends in genetic analysis. London: Oxford University Press 1958
Pritchard, R.H.: The linear arrangement of a series of alleles of Aspergillus nidulans. Heredity 9, 343–371 (1955)
Roman, H.L.: Studies of gene mutation in Saccharomyces. Cold Spr. Harb. Symp. quant. Biol. 21, 175–183 (1956)
Roman, H.L., Jacob, F.: A comparison of spontaneous and ultraviolet induced allelic recombination with reference to the recombination of outside markers. Cold Spr. Harb. Symp. quant. Biol. 23, 155–160 (1958)
Ruddle, F.H.: Linkage analysis in man by somatic cell genetics. Nature (Lond.) 242, 165–169 (1973)
Saltzmann, J., DeMars, R., Benke, P.: Single allele expression at an X-linked hyperuricemia locus in heterozygous human cells. Proc. nat. Acad. Sci. (Wash.) 60, 545–552 (1968)
Sekiguchi, T., Sekiguchi, F.: Interallelic complementation in hybrid cells derived from Chinese hamster diploid clones deficient in hypoxanthine-guanine phosphoribosyltransferase activity. Exp. Cell Res. 77, 391–403 (1973)
Sekiguchi, T., Sekiguchi, F., Tomii, S.: Genetic complementation in hybrid cells derived from mutagen-induced mouse clones deficient in HGPRT activity. Exp. Cell Res. 93, 207–218 (1975)
Shaw, M.W., Cohen, M.M.: Chromosome exchanges in human leukocytes induced by mitomycin-C. Genetics 51, 181–190 (1965)
Siminovitch, L.: On the nature of hereditable variation in culture somatic cells. Cell 7, 1–11 (1976)
Subak-Sharpe, H., Burk, R.R., Pitts, J.D.: Metabolic co-operation between biochemically marked mammalian cells in tissue culture. J. Cell Sci. 4, 353–367 (1969)
Thompson, L.H., Harkins, J.L., Stanners, C.P.: A mammalian cell mutant with a temperature-sensitive leucyl transfer RNA synthetase. Proc. nat. Acad. Sci. (Wash.) 70, 3094–3098 (1973)
Williams, J.F., Young, C.S.H., Austin, P.E.: Genetic analysis of human adenovirus type 5 in permissive and non-permissive cells. Cold Spr. Harb. Symp. quant. Biol. 39, 427–437 (1974)
Zeeland, A.A. van, Diggelen, M.C.E. van, Simons, J.W.I.M.: The role of metabolic co-operation in selection of hypoxanthineguanine phosphoribosyltransferase (HGPRT)-deficient mutants from diploid mammalian cell strains. Mutation Res. 14, 355–363 (1972)
Zimmermann, F.K.: Genetic effects of polynuclear hydrocarbons: Induction of mitotic gene conversion. Z. Krebsforsch. 72, 65–71 (1969)
Author information
Authors and Affiliations
Additional information
Communicated by B.A. Bridges
Rights and permissions
About this article
Cite this article
Tarrant, G.M., Holliday, R. A search for allelic recombination in Chinese hamster cell hybrids. Molec. Gen. Genet. 156, 273–279 (1977). https://doi.org/10.1007/BF00267182
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00267182