Plasmid Isolation

  • Stephen A. Boffey


Some important bacterial genes are not located in the main chromosomal DNA but in independently replicating molecules of circular, double-stranded, ‘plasmid’ DNA. Such genes include those for antibiotic resistance, antibiotic synthesis, toxin production, nitrogen fixation, production of degradative enzymes, and conjugation; so plasmids are obviously of interest in their own right. However, in the context of this book, plasmids are mainly of interest as ‘vectors’ for the cloning of DNA molecules. As will be seen in chapters 11 and 12, it is possible to obtain large quantities of a particular DNA (e.g. cDNA — see Chapter 9) by inserting it into plasmid DNA, to give enlarged plasmids (‘vector’ plasmids) which can be introduced into a suitable host bacterium in which they will replicate. Culture of the cells will result in the production of more plasmid DNA, which can then be isolated from the cells and the inserted DNA recovered.


Buoyant Density Plasmid Isolation Vertical Rotor Preparative Electrophoresis Nucleic Acid Preparation 
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  1. 1.
    F. Bolivar and K. Backman, ‘Plasmids of Escherichia colias cloning vectors’, Methods in Enzymology, 68 (1979), 245–67.PubMedCrossRefGoogle Scholar
  2. 2.
    M. Kahn, R. Kolter, C. Thomas, D. Figurski, R. Meyer, E. Remaut and D.R. Helinski, ‘Plasmid cloning vehicles derived from plasmids ColE1, F, R6K, and RK2’, Methods in Enzymology, 68 (1979), 268–80.PubMedCrossRefGoogle Scholar
  3. 3.
    R.W. Old and S.B. Primrose, Principles of gene manipulation. 2nd edn. Studies in microbiology, 2 (Blackwell, Oxford/University of California Press, Berkeley and Los Angeles, 1981), 33.Google Scholar
  4. 4.
    D.B. Clewell and D.R. Helinski, ‘Properties of a supercoiled deoxyribonucleic acid-protein relaxation complex and strand specificty of the relaxation event’, Biochemistry, 9 (1970), 4428–40.PubMedCrossRefGoogle Scholar
  5. 5.
    M.V. Norgard, ‘Rapid and simple removal of contaminating RNA from plasmid DNA without the use of RNase’, Anal. Biochem., 113 (1981), 34–42.PubMedCrossRefGoogle Scholar
  6. 6.
    H.C. Birnboim and J. Doly, ‘A rapid alkaline extraction procedure for screening recombinant plasmid DNA’, Nucleic Acids Res., 7 (1979), 1513–23.PubMedCrossRefGoogle Scholar
  7. 7.
    D.S. Holmes and M. Quigley, A rapid boiling method for the preparation of bacterial plasmids. Anal. Biochem., 114 (1981), 193–7.PubMedCrossRefGoogle Scholar
  8. 8.
    E. Layne, ‘Spectrophotometic and turbidimetric methods for measuring proteins’, Methods in Enzymology, 3 (1957), 447–54.CrossRefGoogle Scholar
  9. 9.
    Birnboim and Doly, ‘Rapid alkaline extraction’.Google Scholar
  10. 10.
    Norgard, ‘Rapid and simple removal of contaminating RNA’.Google Scholar
  11. 11.
    R.F. Schleif and P.C. Wensink, Practical methods in molecular Biology (Springer-Verlag, New York, 1981).CrossRefGoogle Scholar
  12. 12.
    Norgard, ‘Rapid and simple removal of contaminating RNA’.Google Scholar
  13. 13.
    J.R. Wells and C.F. Brunk, ‘Rapid CsCl gradients using a vertical rotor’, Anal. Biochem., 97 (1979), 196–201.PubMedCrossRefGoogle Scholar
  14. 14.
    M. Fennewald, W. Prevatt, R. Meyer and J. Shapiro, ‘Isolation of Inc P-2 plasmid DNA from Pseudomonas aeruginosa’, Plasmid, 1 (1978), 164–73.PubMedCrossRefGoogle Scholar
  15. 15.
    M. Shoyab and A. Sen, ‘The isolation of extrachromosomal DNA by hydroxyapatite chromatography’, Methods in Enzymology, 68 (1979), 199–206.PubMedCrossRefGoogle Scholar
  16. 16.
    BDH, Poole, England, ‘Hydroxyapatite for nucleic acid work’, Applications pamphlet 1227MP/5.0/1079.Google Scholar
  17. 17.
    LKB, Bromma, Sweden, ‘HA-Ultrogel hydroxyapatite-agarose gel for adsorption chromatography’, Instruction Manual.Google Scholar
  18. 18.
    R.L. Pearson, J.F. Weissand A.D. Kelmers, ‘Improved separation of transfer RNAs on polychlorotrifluoroethylene-supported reversed-phase chromatography columns’, Biochim. Biophys. Acta, 228 (1971), 770–4.PubMedGoogle Scholar
  19. 19.
    R.D. Wells, S.C. Hardies, G.T. Horn, B. Klein, J.E. Larson, S.K. Neuendorf, N. Panayotatos, R.K. Patient and E. Selsing, Methods in Enzymology, 65 (1980), 327–47.PubMedCrossRefGoogle Scholar
  20. 20.
    Bethesda Research Laboratories Inc., Gaithersburg, USA, BRL Catalogue (1981), 86.Google Scholar

Copyright information

© John M. Walker and Wim Gaastra 1983

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  • Stephen A. Boffey

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