Summary
Amber mutants of the mini-F plasmid pML31 were isolated with the mutagen hydroxylamine. Under non-permissive conditions amber mutants segregate and show no incorporation of label into supercoiled plasmid DNA in double-label experiments. Wild-type and one mutant of mini-F were integrated by recombinant DNA techniques into the single EcoRI site of plasmid vector pBR322. Plasmid specific proteins were labeled in minicells and analysed by SDS-PAGE. A 34,000 dalton molecular weight protein was identified to be missing in the amber mutant of plasmid mini-F.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Adler HJ, Fisher WD, Cohen A, Hardigree AA (1966) Miniature Escherichia coli cells deficient in DNA. Proc Natl Acad Sci USA 57:32–326
Andrés I, Slocombe PM, Cabello F, Timmis JK, Lurz R, Burkhardt HJ, Timmis KN (1979) Plasmid replication functions. II. Cloning analysis of the RepA replication region of antibiotic resistance plasmid R6-5. Mol Gen Genet 168:1–25
Bachmann BJ (1972) Pedigrees of some mutant strains of Escherichia coli K-12. Bacteriol Rev 36:525–557
Bolivar F, Rodriguez RL, Greene PJ, Betlach MC, Heyneker HL, Boyer HW (1977) Construction and characterization of new cloning vehicles. II A multipurpose cloning system. Gene 2:95–113
Bonner WM, Laskey RA (1974) A film detection method for tritium-labeled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem 46:83–88
Collins J, Yanofsky S, Helinski DR (1978) Involvement of the DNA-protein complex in the replication of plasmid ColEl. Mol Gen Genet 167:21–28
Cuzin F, Jacob F (1967) Mutations de l'épisome F d'Escherichia coli K12. II. Mutants à replication thermosensible. Ann Inst Pasteur Paris 112:398–418
Deleuve G, Cabezón T, Ghysen A, Herzog A, Bollen A (1977) Amber mutations in Escherichia coli essential genes: Isolation of mutants affected in the ribosome. Mol Gen Genet 157:149–153
Dougan G, Sherratt D (1977) The transposon Tn1 as a probe for studying ColEl structure and function. Mol Gen Genet 151:151–160
Eichenlaub R, Figurski D, Helinski DR (1977) Bidirectional replication from a unique origin in a mini-F plasmid. Proc Natl Acad Sci USA 74:1138–1141
Eichenlaub R (1979) Mutants of the mini-F plasmid pML31 thermosensitive in replication. J Bacteriol 138:559–566
Figurski D, Kolter R, Meyer R, Kahn M, Eichenlaub R, Helinski DR (1978) Replication regions of plasmids ColE1, F, R6K, and RK2. In: Schlessinger D (ed) Microbiology-1978. Am Soc Micro Washington DC, p 105
Gustafsson P, Nordström K (1978) Temperature-dependent and amber copy mutants of plasmid Rldrd-19 in Escherichia coli. Plasmid 1:134–144
Hashimoto-Gotoh T, Sekiguchi M (1977) Mutations to temperature sensitivity in R plasmid pSC101. J Bacteriol 131:404–412
Hashimoto T, Sekiguchi M (1976) Isolation of temperature-sensitive mutants of R plasmids by in vitro mutagenesis with hydroxylamine. J Bacteriol 127:1561–1563
Humphreys GO, Willshaw GA, Smith HR, Anderson ES (1976) Mutagenesis of plasmid DNA with hydroxylamine: isolation of mutants of multi-copy plasmids. Mol Gen Genet 145:101–108
Inuzuka M, Helinski DR (1978) Requirement of a plasmid encoded protein for the replication in vitro of plasmid R6K. Proc Natl Acad Sci USA 75:5381–5385
Kennedy N, Bentin L, Achtmann M, Skurray R, Rahmdsorf U, Herrlich P (1977) Conjugation proteins encoded by the F sex factor. Nature 270:580–585
Kollek R, Oertel W, Goebel W (1978) Isolation and characterization of the minimal fragment required for autonomous replication (“basic replicon”) of a copy mutant (pKN102) of the antibiotic resistance factor R1. Mol Gen Genet 162:51–57
Laskey RA, Mills AD (1975) Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem 56:335–341
Lugtenberg B, Meijers J, Peters R, van der Hock P, van Alphen L (1975) Electrophoretic resolution of the “major outer membrane protein” of Escherichia coli K12 into four bands. FEBS Lett 58:254–258
Manis J, Kline BC (1978) F plasmid incompatibility and copy number genes: their map locations and interaction. Plasmid 1:492–507
Novick R, Wyman L, Bouanchaud D, Murphy E (1978) Plasmid life cycle in Staphylococcus aureus. In: Schlessinger D (ed) Microbiology-1974. Am Soc Micro Washington DC, p 115
Smith DJ, Blattner FR, Davies J (1976) The isolation and partial characterization of a new restriction endonuclease from Providencia stuartii. Nucleic Acids Res 3:343–353
Wehlmann H, Eichenlaub R (1979) Cloning of restriction fragments of plasmid mini-F: mapping of replication and incompatibility regions and characterization of RNA and proteins. Hoppe Seyler's Z Physiol Chem 360:1047–1048
Yamamoto R, Kaji A (1977) Replication of thermosensitive Rts1 plasmid deoxyribonucleic acid at the nonpermissive temperature. J Bacteriol 132:90–99
Author information
Authors and Affiliations
Additional information
Communicated by J. Schell
Rights and permissions
About this article
Cite this article
Eichenlaub, R., Wehlmann, H. Amber-mutants of plasmid mini-F defective in replication. Molec. Gen. Genet. 180, 201–204 (1980). https://doi.org/10.1007/BF00267370
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00267370