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A method for the purification ofE. coli plasmid DNA by homogeneous lysis and polyethylene glycol precipitation

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Abstract

A procedure is described for the isolation and purification ofE. coli plasmid DNA by polyethylene glycol precipitation. The method is rapid, simple, inexpensive and amenable to both small and large scale manipulation. This procedure involves lysis of bacterial cells by treatment with pronase in sodium dodecyl sulfate, removal of chromosomal DNA by centrifugation, precipitation of residual nucleic acids with polyethylene glycol and removal of RNA by precipitation with LiCl. Plasmid DNA purified as described is pure enough for restriction endonuclease analysis, for use as a vector for the cloning of cDNA or synthetic DNA, or for use as a template in anE. coli transcription-translation cell-free system.

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Abbreviations

PEG:

polyethylene glycol

TE buffer:

10 mM Tris-HCl pH 7.5, 1 mM EDTA

SDS:

sodium dodecyl sulfate

tet:

tetracycline

amp:

ampicillin

kan:

kanamycin

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Author notes

  1. R. Glick

    Present address: Biology Department, University of Waterloo, N2L 3G1, Waterloo, Ontario, Canada

Authors and Affiliations

  1. Department of Biochemistry, University of Toronto, M5S 1A8, Toronto, Ontario, Canada

    D. Pulleyblank

  2. Molecular Genetics and Biochemistry Group, BIO LOGICALS, 7 Hinton Avenue North, K1Y 4P1, Ottawa, Ontario, Canada

    M. Michalak, St. Laurent Daisley & R. Glick

Authors
  1. D. Pulleyblank
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  2. M. Michalak
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  3. St. Laurent Daisley
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  4. R. Glick
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Pulleyblank, D., Michalak, M., Laurent Daisley, S. et al. A method for the purification ofE. coli plasmid DNA by homogeneous lysis and polyethylene glycol precipitation. Mol Biol Rep 9, 191–195 (1983). https://doi.org/10.1007/BF00775367

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  • DOI: https://doi.org/10.1007/BF00775367

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