Hostname: page-component-745bb68f8f-grxwn Total loading time: 0 Render date: 2025-01-13T01:38:57.107Z Has data issue: false hasContentIssue false
  • English
  • Français

The role of oriT in tra-dependent enhanced recombination between mini-F-lac-oriT and λplac5

Published online by Cambridge University Press:  14 April 2009

Jeffrey R. Carter ,
Dipty R. Patel  and
Ronald D. Porter
Jeffrey R. Carter
Affiliation:
Department of Molecular and Cell Biology, S-101 Frear Laboratory, The Pennsylvania State University, University Park, PA 16802, USA
Dipty R. Patel
Affiliation:
Department of Molecular and Cell Biology, S-101 Frear Laboratory, The Pennsylvania State University, University Park, PA 16802, USA
Ronald D. Porter*
Affiliation:
Department of Molecular and Cell Biology, S-101 Frear Laboratory, The Pennsylvania State University, University Park, PA 16802, USA
*
† Corresponding author.
Rights & Permissions [Opens in a new window]

Summary

Core share and HTML view are not available for this content. However, as you have access to this content, a full PDF is available via the ‘Save PDF’ action button.

Recombination between F42lac and λplac5 is typically 20- to 50-fold more efficient than recombination between chromosomal lac and λplac5. This enhancement of recombination is recBCD-dependent and requires the expression of genes from the tra regulon of the F factor. Also required is oriT, the origin of F factor conjugational transfer, which must be located in-cis to the cellular copy of lac. In this study we show that enhanced recombination is not supported by an oriT point mutant that reduces oriT function in conjugation. We also present evidence that the activation of oriT for recombination enhancement involves the same strand-specific nick that is required for conjugal DNA transfer. Although it is thought that the role of oriT in recombination enhancement is related to the facilitated entry of RecBCD enzyme into the DNA duplex, we were unable to detect any double-strand breakage at oriT.

Type
Research Article
Information
Genetics Research , Volume 59 , Issue 3 , June 1992 , pp. 157 - 165
Copyright
Copyright © Cambridge University Press 1992

References

Bachmann, B. (1972). Pedigrees of some mutant strains of Escherichia coli K-12. Bacteriological Reviews 36, 525–557.Google Scholar
Birge, E. A. & Low, K. B. (1974). Detection of transcribable recombination products following conjugation in Rec+, RecB− and RecC− strains of Escherichia coli K-12. Journal of Molecular Biology 83, 447–457.Google Scholar
Birnboim, H. C. & Doly, J. (1979). A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Research 7, 1513–1523.Google Scholar
Carter, J. R. & Porter, R. D. (1991). traY and tral are required for oriT-dependent enhanced recombination between lac-containing plasmids and λplac5. Journal of Bacteriology 173, 1027–1034.Google Scholar
Chaconas, G.de Bruijn, F. J.Casadaban, M. J.Lupski, J. R.Kwoh, T. J.Harshey, R. M.Dubow, M. S. & Bukhari, A. I. (1981). In vitro and in vivo manipulations of bacteriophage Mu DNA: cloning of Mu ends and construction of mini-Mu's carrying selectable markers. Gene 13, 37–46.Google Scholar
Clark, J. M. (1988). Novel non-templated nucleotide addition reactions catalyzed by procaryotic and eucaryotic DNA polymerases. Nucleic Acids Research 16, 9677–9686.Google Scholar
Everett, R. & Willetts, N. (1980). Characterization of an in vivo system for nicking at the origin of conjugal DNA transfer of the sex factor F. Journal of Molecular Biology 136, 129–150.Google Scholar
Everett, R. & Willetts, N. (1982). Cloning, mutation, and location of the F origin of conjugal transfer. EMBO Journal 1, 747–753.CrossRefGoogle Scholar
Hattori, M. & Sakaki, Y. (1986). Dideoxy sequencing method using denatured plasmid templates. Analytical Biochemistry 152, 232–238.Google Scholar
Kushner, S. R. (1978). An improved method for transformation of Escherichia coli with ColE1 derived plasmids, pp. 17–23. In Genetic Engineering (ed. Boyer, H. B. and Nicosia, S.). Amsterdam: Elsevier/North-Holland Publishing Co.Google Scholar
Low, B. (1973). Rapid mapping of conditional and auxotrophic mutations in Escherichia coli K-12. Journal of Bacteriology 113, 798–812.Google Scholar
Low, B. & Wood, T. H. (1965). A quick and efficient method for interruption of bacterial conjugation. Genetical Research 6, 300–303.CrossRefGoogle ScholarPubMed
Maniatis, T.Frisch, E. F. & Sambrook, J. (1982). Molecular Cloning: a Laboratory Manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press.Google Scholar
Manis, J. J. & Kline, B. (1977). Restriction endonuclease mapping and mutagenesis of the F sex factor replication region. Molecular and General Genetics 152, 175–182.Google Scholar
Miller, J. H. (1972). Experiments in Molecular Genetics. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press.Google Scholar
Porter, R. D. (1981). Enhanced recombination between F42lac and λplac5: Dependence on F42lac fertility functions. Molecular and General Genetics 184, 355–358.Google Scholar
Porter, R. D. (1982). Recombination properties of P1dlac. Journal of Bacteriology 152, 345–350.Google Scholar
Porter, R. D. (1983). Specialized transduction with λplac5: involvement of the RecE and RecF recombination pathways. Genetics 105, 247–257.CrossRefGoogle Scholar
Porter, R. D.Lark, M. W. & Low, K. B. (1981). Specialized transduction with λplac5: Dependence on recA and on configuration of lac and attλ. Journal of Virology 38, 497–503.CrossRefGoogle Scholar
Porter, R. D.McLaughlin, T. & Low, B. (1978). Transduction versus ‘conjuduction’: Evidence for multiple roles for exononuclease V in genetic recombination in Escherichia coli. Cold Spring Harbor Symposia on Quantitative Biology 43, 1043–1047.Google Scholar
Porter, R. D.Welliver, R. A. & Witkowski, T. A. (1982). Specialized transduction with λplac5: Dependence on recB. Journal of Bacteriology 150, 1485–1488.CrossRefGoogle Scholar
Sanger, F.Nicklen, S.Coulson, A. R. (1977). DNA sequencing with chain terminating inhibitors. Proceedings of the National Academy of Sciences of USA 74, 5463–5467.Google Scholar
Seifert, H. S. (1984). Ph.D. thesis. The Pennsylvania State University, University Park, PA, USA.Google Scholar
Seifert, H. S. & Porter, R. D. (1984). Enhanced recombination between λplac5 and F42lac: Identification of cisand trans-acting factors. Proceedings of the National Academy of Sciences USA 81, 7500–7504.CrossRefGoogle Scholar
Smith, G. R. (1988). Homologous recombination in procaryotes. Microbiological Reviews 52, 1–28.Google Scholar
Southern, E. M. (1975). Detection of specific sequences among DNA fragments separated by gel electrophoresis. Journal of Molecular Biology 98, 503–517.Google Scholar
Taylor, A. F. & Smith, G. R. (1985). Substrate specificity of the DNA unwinding activity of the RecBC enzyme of Escherichia coli. Journal of Molecular Biology 185, 431–43.Google Scholar
Thompson, T. L.Centola, M. B. & Deonier, R. C. (1989). Location of the nick at oriT of the F plasmid. Journal of Molecular Biology 207, 505–512.Google Scholar
Traxler, B. A. & Minkley, E.G. (1987). Revised genetic map of the distal end of the F transfer operon: Implications for DNA Helicase I, nicking at oriT and conjugal DNA transport. Journal of Bacteriology 169, 3251–3259.Google Scholar
Wehlmann, H. & Eichenlaub, R. (1980). Plasmid mini-F encoded. Molecular and General Genetics 180, 205–211.Google Scholar
Yancey, S. D. & Porter, R. D. (1985). General recombination in E. coli K-12: in vivo role of RecBC enzyme. Journal of Bacteriology 162, 29–34.Google Scholar