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Biological assay of prokaryotic genes in mouse cells following DNA mediated transformation

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Summary

Genes coding for leucine biosynthesis in Bacillus subtilis were introduced into mouse LTK- cells by co-transformation with thymidine kinase+ (tk) DNA. Genomic DNA from the tk+ transformants was used to transform competent cultures of different B. subtilis leucine auxotrophs. Each auxotroph was transformed to prototrophy at a similar frequency and the number of leucine gene sequences per transformant genome as deduced by the B. subtilis bioassay strongly correlated with the number estimated by hybridization methods. Tk- subclones were obtained by plating the transformants in 5′-bromodeoxyuridine. One subclone still contained the non-selected leucine gene sequences and could transform auxotrophs of B. subtilis. No deletions or rearrangements in the linkage relationships of the leucine genes occurred in the LTK- cells that inhibited transformation of B. subtilis.

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References

  • Benton DW, Davis RW (1977) Screening λgt recombinant clones by hybridization to single plaques in situ. Science 196:180–181

    Google Scholar 

  • Bodmer WF, Ganesan AT (1964) Biochemical and genetic studies of integration and recombination in Bacillus subtilis transformation. Genetics 50:717–738

    Google Scholar 

  • Breathnach R, Mantei N, Chambon P (1980) Correct splicing of a chicken ovalbumin gene transcript in mouse L cells. Proc Natl Acad Sci USA 77:740–744

    Google Scholar 

  • Burton K (1956) A study of conditions and mechanisms of diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem J 62:315–323

    Google Scholar 

  • Chi NW, Ehrlich SD, Lederberg J (1978) Functional expression of two Bacillus subtilis chromosomal genes in Escherichia coli. J Bacteriol 133:816–821

    Google Scholar 

  • Clewell D, Helinski DR (1972) Effect of growth conditions on the formation of the relaxation complex of supercoiled ColEl deoxyribonucleic acid and protein in Escherichia coli. J Bacteriol 110:1135–1146

    Google Scholar 

  • Duncan GH, Wilson GA, Young FE (1978) Mechanism of integrating foreign DNA during transformation of Bacillus subtilis. Proc Natl Acad Sci USA 75:3664–3668

    Google Scholar 

  • Enquist LW, Would GFV, Wagner M, Smiley JR, Summers WC (1979) Construction and characterization of a recombinant plasmid encoding the gene for the thymidine kinase of Herpes simplex type 1 virus. Gene 7:335–342

    Google Scholar 

  • Graham FL, van der Eb AJ (1973) A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52:456–467

    Google Scholar 

  • Harris-Warrick RM, Elkana Y, Ehrlich SD, Lederberg J (1975) Electrophoretic separation of Bacillus subtilis genes. Proc Natl Acad Sci USA 72:2207–2211

    Google Scholar 

  • Helling RB, Goodman HM, Boyer HW (1974) Analysis of endonuclease R·EcoRl fragments of DNA from lambdoid bacteriophages and other viruses by agarose-gel electrophoresis. J Virol 14:1235–1244

    Google Scholar 

  • Henner DJ, Hoch JA (1980) The Bacillus subtilis chromosome. Microb Rev 44:57–82

    Google Scholar 

  • Jackson EN, Miller HI, Adams ML (1978) EcoRl restriction endonuclease cleavage site map of bacteriophage P22 DNA. J Mol Biol 118:347–363

    Google Scholar 

  • Keggins KM, Lovett PS, Duvall EJ (1978) Molecular cloning of genetically active fragments of Bacillus DNA in Bacillus subtilis and properties of the vector plasmid pUB110. Proc Natl Acad Sci USA 75:1423–1427

    Google Scholar 

  • La EC, Woo SLC, Bordelon-Riser ME, Fraser TH, O'Malley BW (1980) Ovalbumin is synthesised in mouse cells transformed with the natural chicken ovalbumin gene. Proc Natl Acad Sci USA 77:244–248

    Google Scholar 

  • Littlefield JW (1964) Selection of hybrids from matings of fibroblasts in vitro and their presumed recombinants. Science 145:709–710

    Google Scholar 

  • Nagahari K, Sakaguchi K (1978) Cloning of Bacillus subtilis leucine A, B and C genes with Escherichia coli plasmids and expression of the leuC gene in E. coli. Mol Gen Genet 158:263–270

    Google Scholar 

  • Nunberg JH, Kaufman RJ, Chang ACY, Cohen SN, Schimke RT (1980) Structure and genomic organization of the mouse dihydrofolate reductase gene. Cell 19:355–364

    Google Scholar 

  • Pellicer A, Robins D, Wold B, Sweet R, Jackson J, Lowy I, Roberts JM, Sim GK, Silverstein S, Axel R (1980) Altered genotype and phenotype by DNA-mediated gene transfer. Science 209:1414–1422

    Google Scholar 

  • Pellicer A, Wigler M, Axel R, Silverstein S (1978) The transfer and stable integration of the HSV thymidine kinase gene into mouse cells. Cell 14:133–141

    Google Scholar 

  • Perucho M, Hanahan D, Lipsich L, Wigler M (1980) Isolation of the chicken thymidine kinase gene by plasmid rescue. Nature 285:207–210

    Google Scholar 

  • Perucho M, Hanahan D, Wigler M (1980) Genetic and physical linkage of exogenous sequences in transformed cells. Cell 22:309–317b

    Google Scholar 

  • Rigby PWJ, Dieckman M, Rhodes C, Berg P (1977) Labeling deoxyribonucleic acid to a high specific activity in vitro by knick translation with DNA polymerase 1. J Mol Biol 113:237–251

    Google Scholar 

  • Sharp PA, Sugden B, Sambrook J (1973) Detection of two restriction endonuclease activities in Haemophilus parainfluenza using analytical agarose- ethidium bromide electrophoresis. Biochemistry 12:3055–3063

    Google Scholar 

  • Southern EM (1975) Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517

    Google Scholar 

  • Southern E (1979) Gel electrophoresis of restriction fragments. In: Wu R (ed) Methods in enzymology, vol 68. Academic Press, New York, p 152–176

    Google Scholar 

  • Spizizen J (1958) Transformation of biochemically deficient strains of Bacillus subtilis by deoxyribonucleate. Proc Natl Acad Sci USA 44:1072–1078

    Google Scholar 

  • Stewart C (1969) Physical heterogeneity among Bacillus subtilis deoxyribonucleic acid molecules carrying particular genetic markers. J Bacteriol 98:1239–1247

    Google Scholar 

  • Wahl GM, Stern M, Stark GR (1979) Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate. Proc Natl Acad Sci USA 76:3683–3687

    Google Scholar 

  • Ward JB, Zahler SA (1973) Genetic studies of leucine biosynthesis in Bacillus subtilis. J Bacteriol 116:719–726

    Google Scholar 

  • Wigler M, Pellicer A, Silverstein S, Axel R (1978) Biochemical transfer of single- copy genes using total cellular DNA as donor. Cell 14:725–731

    Google Scholar 

  • Wigler M, Pellicer A, Silverstein S, Axel R, Urlaub G, Chasin L (1979) DNA-mediated transfer of the adenine phosphoribosyltransferase locus into mammalian cells. Proc Natl Acad Sci USA 76:1373–1376

    Google Scholar 

  • Wigler M, Sweet R, Sim GK, Wold B, Pellicer A, Lacy E, Maniatis T, Silverstein S, Axel R (1979) Transformation of mammalian cells with genes from prokaryotes and eukaryotes. Cell 16:777–785

    Google Scholar 

  • Willecke K, Klomfass M, Micrau R, Döhmer J (1979) Intraspecies transfer via total cellular DNA of the gene for hypoxanthine phosphoribosyltransferase into cultured mouse cells. Mol Gen Genet 170:179–185

    Google Scholar 

  • Wold B, Wigler M, Lacy E, Maniatis T, Silverstein S, Axel R (1979) Introduction and expression of a rabbit β-globin gene in mouse fibroblasts. Proc Natl Acad Sci USA 76:5684–5688

    Google Scholar 

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Authors and Affiliations

  1. Department of Genetics, Lt., J. P. Kennedy Laboratories, Stanford Medical School, 94305, Stanford, CA, USA

    John I. Yoder & A. T. Ganesan

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  1. John I. Yoder
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  2. A. T. Ganesan
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Communicated by M.M. Green

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Yoder, J.I., Ganesan, A.T. Biological assay of prokaryotic genes in mouse cells following DNA mediated transformation. Molec. Gen. Genet. 181, 525–531 (1981). https://doi.org/10.1007/BF00428747

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  • DOI: https://doi.org/10.1007/BF00428747

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