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Expression of a neomycin phosphotransferase gene from Streptomyces fradiae in Escherichia coli after interplasmidic recombination

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Summary

Plasmid pIJ2 carrying the neomycin phosphotransferase gene of Streptomyces fradiae was fused to E. coli plasmid pBR325 and the hybrid molecules were introduced into E. coli K12 by transformation. The neomycin phosphotransferase gene of the hybrid plasmid was not expressed in E. coli, except after interplasmidic recombination. Physical analysis of such an in vivo recombinant plasmid revealed that the recombination brought one neomycin phosphotransferase gene to a position downstream from the tet-promoter of pBR325. Subcloning experiments indicated that this is the gene copy expressed, and that transcription is initiated at the tet-promoter of pBR325.

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References

  • Beck E, Somer R, Auerswald AE, Kurz C, Zink B, Osterburg G, Schaller H, Sugimoto K, Sugisaki H, Okamoto T, Takanami M (1978) Nucleotide sequence of bacteriophage fd DNA. Nucl Acids Res 5:4495–4503

    Google Scholar 

  • Bibb MJ, Freeman RF, Hopwood DA (1977) Physical and genetical characterisation of a second sex factor, SCP2, for Streptomyces coelicolor A3(2). Mol Gen Genet 154:155–166

    Google Scholar 

  • Bibb MJ, Ward JM, Hopwood DA (1978) Transformation of plasmid DNA into Streptomyces at high frequency. Nature 274:398–400

    Google Scholar 

  • Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucl Acids Res 7:1513–1523

    Google Scholar 

  • Bolivar F (1978) Construction and characterization of new cloning vehicles. III. Derivatives of plasmid pBR322 carrying unique EcoRI sites for selection of EcoRI generated recombinant DNA molecules. Gene 4:121–136

    Google Scholar 

  • Bolivar F, Rodriguez RL, Greene PJ, Betlach MC, Heyneker HL, Boyer Hw, Crosa JH, Falkow S (1977) Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene 2:95–113

    Google Scholar 

  • Boyer HW, Roulland-Dussoix D (1969) A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol 41:459–472

    Google Scholar 

  • Chang ACY, Cohen SN (1974) Genome construction between bacterial species in vitro: replication and expression of Staphylococcus plasmid genes in Escherichia coli. Proc Natl Acad Sci USA 71:1030–1034

    Google Scholar 

  • Chater KF, Hopwood DA, Kieser T, Thompson CJ (1982) Gene cloning in Streptomyces. Curr Topics Microbiol Immunol 96:69–95

    Google Scholar 

  • Cohen SN, Chang ACY, Asu L (1972) Nonchromosomal antibiotic resistance in bacteria: genetic transformation of Escherichia coli by R-factor DNA. Proc Natl Acad Sci USA 69:2110–2114

    Google Scholar 

  • Courvalin P, Fiandt M (1980) Aminoglycoside-modifying enzymes of Staphylococcus aureus: expression in Escherichia coli. Gene 9:247–269

    Google Scholar 

  • Courvalin P, Weisblum B, Davies J (1977) Aminoglycoside-modifying enzymes of an antibiotic-producing bacterium acts as a determinant of antibiotic resistance in Escherichia coli. Proc Natl Acad Sci USA 74:999–1003

    Google Scholar 

  • Davis RW, Simon M, Davidson N (1971) Electron microscope heteroduplex methods for mapping regions of base sequence homology in nucleic acids. Methods Enzymol 21:413–428

    Google Scholar 

  • Ehrlich SD, Bursztyn-Pettegrew H, Stroynowski I, Lederberg J (1976) Expression of the thymidylate synthetase gene of Bacillus subtilis bacteriophage Phi-3-T in Escherichia coli. Proc Natl Acad Sci USA 73:4145–4149

    Google Scholar 

  • Haas MJ, Dowding JE (1975) Aminoglycoside-modifying enzymes. Methods Enzymol 43:611–628

    Google Scholar 

  • Horinouchi S, Uozumi T, Beppu T (1980) Cloning of Streptomyces DNA into Escherichia coli: absence of heterospecific gene expression of Streptomyces genes in E. coli. Agric Biol Chem 44:367–381

    Google Scholar 

  • Iida S, Meyer J, Arber W (1981) Cointegrates between bacteriophage P1 DNA and plasmid pBR322 derivatives suggest molecular mechanism for P1-mediated transduction of small plasmids. Mol Gen Genet 184:1–10

    Google Scholar 

  • Ikeda H, Morya K, Matsumoto T (1981) In vitro study of illegitimate recombination: involvement of DNA gyrase. Cold Spring Harbor Symp Quant Biol 45:399–408

    Google Scholar 

  • Kilbane JJ, Malamy MH (1980) F factor mobilization of nonconjugative chimeric plasmids in Escherichia coli: general mechanisms and role for site-specific recA-independent recombination at ori VI. J Mol Biol 143:73–93

    Google Scholar 

  • Lennox ES (1955) Transduction of linked genetic characters of the host by bacteriophage P1. Virology 1:190–206

    Google Scholar 

  • Lomovskaya ND, Mrktumian NM, Gostimskaya NL, Danilenko VN (1972) Characterization of temperate actinophage πC31 isolated from Streptomyces coelicolor A3(2). J Virol 9:258–262

    Google Scholar 

  • Mahler I, Halvorson HO (1977) Transformation of Escherichia coli and Bacillus subtilis with a hybrid plasmid molecule. J Bacteriol 131:374–377

    Google Scholar 

  • Masters M, Andresdottir V, Wolf-Walz H (1979) Plasmids carrying oriC can integrate at or near the chromosome origin of Escherichia coli in the absence of a functional recA product. Cold Spring Harbor Symp Quant Biol 43:1069–1072

    Google Scholar 

  • Pogue-Geile KL, Dassarma S, King SR, Jaskunas SR (1980) Recombination between bacteriophage lambda and plasmid pB322 in Escherichia coli. J Bacteriol 142:992–1003

    Google Scholar 

  • Prentki P, Karch F, Iida S, Meyer J (1981) The plasmid cloning vector pBR325 contains a 482 base-pair-long inverted duplication. Gene 14:289–299

    Google Scholar 

  • Robbins PW, Wirth DF, Hering C (1981) Expression of the Streptomyces enzyme endoglycosidase H in Escherichia coli. J Biol Chem 256:10640–10644

    Google Scholar 

  • Rodgers WH, Springer W, Young FE (1982) Cloning and expression of a Streptomyces fradiae neomycin resistance gene in Escherichia coli. Gene 18:133–141

    Google Scholar 

  • Rosenberg M, Court D (1979) Regulatory sequences involed in the promotion and termination of RNA transcription. Annu Rev Genet 13:319–353

    Google Scholar 

  • Schottel JL, Bibb MJ, Cohen SN (1981) Cloning and expression in Streptomyces lividans of antibiotic resistance genes derived from Escherichia coli. J Bacteriol 146:360–368

    Google Scholar 

  • Stüber D, Bujard H (1981) Organization of transcriptional signals in plasmids pBR322 and pACYC184. Proc Natl Acad Sci USA 78:167–171

    Google Scholar 

  • Thompson CJ, Ward JM, Hopwood DA (1980) DNA cloning in Streptomyces: resistance genes from antibiotic-producing species. Nature 286:525–529

    Google Scholar 

  • Timmis KN, Cabello F, Cohen SN (1978) Cloning and characterization of EcoRI and HindIII restriction endonuclease-generated fragments of antibiotic resistance plasmids R6-5 and R6. Mol Gen Genet 162:121–137

    Google Scholar 

  • Westmoreland BC, Szybalski W, Ris H (1969) Mapping of deletions and substitutions in heteroduplex DNA molecules of bacteriophage lambda by electron microscopy. Science 163:1343–1348

    Google Scholar 

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Communicated by H. Böhme

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Schupp, T., Toupet, C., Stålhammar-Carlemalm, M. et al. Expression of a neomycin phosphotransferase gene from Streptomyces fradiae in Escherichia coli after interplasmidic recombination. Molec Gen Genet 189, 27–33 (1983). https://doi.org/10.1007/BF00326051

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  • DOI: https://doi.org/10.1007/BF00326051

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