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Plasmid replication functions

II. Cloning analysis of the RepA replication region of antibiotic resistance plasmid R6-5

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Summary

R6-5 is a low copy number, conjugative, FII incompatibility group plasmid that has a molecular length of 102 kb and that specifies resistance against several antibiotics (chloramphenicol, fusidic acid, kanamycin, streptomycin and sulphonamide) and mercury salts. By means of in vitro cloning procedures, mini plasmids have been generated that contain a DNA segment from the essential region of R6-5 that is only 2.6 kb in length. This DNA segment, which consists of two PstI fragments that are adjacent in the parent plasmid, carries all genes and sequences required for the regulated replication and incompatibility properties of R6-5, including its origin of replication, OriV, an essential function that has been designated RepA, and the copy control function, Cop. Three different polypeptides, having monomer molecular weights of 23,000, 10,000 and 9,500 daltons, are synthesized in detectable quantities by minicells carrying pBR322 hybrid plasmids that contain DNA segments from the R6-5 essential region. A spontaneous deletion derivative of a pBR322 hybrid plasmid that carries the R6-5 origin of replication was isolated. Heteroduplex analysis of this derivative plasmid indicates that the deleted DNA segment carries the R6-5 replication origin and that its termini consist of short inverted repeat sequences.

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Author notes

  1. Isabel Andrés

    Present address: Department of Biochemistry, University of Santander, Santander, Spain

  2. Felipe Cabello

    Present address: Department of Microbiology, New York Medical College, Valhalla, New York, USA

Authors and Affiliations

  1. Max-Planck-Institut für Molekulare Genetik, Ihnestraße 63-73, D-1000, Berlin 33

    Isabel Andrés, Patrick M. Slocombe, Joan K. Timmis, Rudolf Lurz & Kenneth N. Timmis

  2. Lehrstuhl für Mikrobiologie der Universität Erlangen-Nürnberg, Erlangen, BRD

    Felipe Cabello & Hans J. Burkardt

Authors
  1. Isabel Andrés
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  2. Patrick M. Slocombe
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  3. Felipe Cabello
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  4. Joan K. Timmis
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  5. Rudolf Lurz
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  6. Hans J. Burkardt
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  7. Kenneth N. Timmis
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Additional information

Communicated by T. Yura

Paper I in this series is Timmis et al., 1978b

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Andrés, I., Slocombe, P.M., Cabello, F. et al. Plasmid replication functions. Molec. Gen. Genet. 168, 1–25 (1979). https://doi.org/10.1007/BF00267929

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