Advertisement

Recombination and Site-Directed Mutagenesis Using Recombination PCR

  • Douglas H. Jones
  • Stanley C. Winistorfer
Part of the Methods in Molecular Biology™ book series (MIMB, volume 67)

Abstract

The polymerase chain reaction (PCR) (1) provides a rapid means for the recombination and site-directed mutagenesis of DNA (2). DNA modification can occur during PCR because the primers are incorporated into the ends of the PCR product. The simplest PCR-based method for site-directed mutagenesis and DNA recombination is recombination PCR.

Keywords

Polymerase Chain Reaction Polymerase Chain Reaction Product Polymerase Chain Reaction Amplification Restriction Endonuclease Digestion Donor Plasmid 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

References

  1. 1.
    Mullis, K., Faloona, F., Scharf, S., Saiki, R., Horn, G., and Erlich, H. (1986) Specific enzymatic amplification of DNA in vitro: the polymerase chain reaction, in Cold Spring Harbor Symposia on Quantitative Biology, vol. LI, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, pp. 263–273.Google Scholar
  2. 2.
    White, B. (1993) Methods in Molecular Biology, vol. 15, PCR Protocols Current Methods and Applications, Humana, Totowa, NJ.Google Scholar
  3. 3.
    Jones, D. H. and Howard, B. H. (1991) A rapid method for recombination and site-specific mutagenesis by placing homologous ends on DNA using polymerase chain reaction. BioTechniques 10, 62–66.PubMedGoogle Scholar
  4. 4.
    Jones, D. H. (1994) PCR mutagenesis and recombination in vivo. PCR Methods Appl. 3, S141–S148.PubMedCrossRefGoogle Scholar
  5. 5.
    Coco, W. M., Rothmel, R. K., Henikoff, S., and Chakrabarty, A. M. (1993) Nucleotide sequence and initial functional characterization of the clcR gene encoding a LysR family activator of the clcABD chlorocatechol operon in Pseudomonas putida. J. Bacteriol. 175, 417–427.PubMedCentralPubMedGoogle Scholar
  6. 6.
    Fridovich-Keil, J. L. and Jinks-Robertson, S. (1993) A yeast expression system for human galactose-1-phosphate uridylyltransferase. Proc. Natl. Acad. Sci. USA 90, 398–402.PubMedCrossRefGoogle Scholar
  7. 7.
    Goulden, M. G., Kohm, B. A, Santa Cruz, S., Kavanagh, T. A., and Baulcombe, D. C. (1993) A feature of the coat protein of potato virus X affects both induced virus resistance in potato and viral fitness. Virology 197, 293–302.PubMedCrossRefGoogle Scholar
  8. 8.
    Gibbs, J. S., Regier, D. A., and Desrosiers, R. C. (1994) Construction and in vitro properties of HIV-1 mutants with deletions in “nonessential” genes. AIDS Res. Hum. Retrovir. 10, 343–350.PubMedCrossRefGoogle Scholar
  9. 9.
    Singh, K. K., Small, G. M., and Lewin, A. S. (1992) Alternative topogenic signals in peroxisomal citrate synthase of Saccharomyces cerevisiae. Mol. Cell. Biol. 12, 5593–5599.PubMedCentralPubMedGoogle Scholar
  10. 10.
    Weiner, M. P., Costa, G. L., Schoettlin, W., Cline, J., Mathur, E., and Bauer, J. C. (1994) Site-directed mutagenesis of double-stranded DNA by the polymerase chain reaction. Gene 151, 119–123.PubMedCrossRefGoogle Scholar
  11. 11.
    Jones, D. H. and Winistorfer, S. C. (1992) Recombinant circle PCR and recombination PCR for site-specific mutagenesis without PCR product purification. BioTechniques 12, 528–534.PubMedGoogle Scholar
  12. 12.
    Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, p. B. 23.Google Scholar
  13. 13.
    Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, p. A. 2.Google Scholar
  14. 14.
    Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, p. A. 4.Google Scholar
  15. 15.
    Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, NY, p. A. 1.Google Scholar
  16. 16.
    Liang, W. and Johnson, J. P. (1988) Rapid plasmid insert amplification with polymerase chain reaction. Nucleic Acids Res. 16, 3579.PubMedCentralPubMedCrossRefGoogle Scholar
  17. 17.
    Lundberg, K. S., Shoemaker, D. D., Adams, M. W. W., Short, J. M., Sorge, J. A., and Mathur, E. J. (1991) High-fidelity amplification using a thermostable DNA polymerase isolated from Pyrococcus furiosus. Gene 108, 1–6.PubMedCrossRefGoogle Scholar
  18. 18.
    Yao, Z., Jones, D. H., and Grose, C. (1992) Site-directed mutagenesis of herpes-virus glycoprotein phosphorylation sites by recombination polymerase chain reaction. PCR Methods Appl. 1, 205–207.PubMedCrossRefGoogle Scholar

Copyright information

© Humana Press Inc. 1997

Authors and Affiliations

  • Douglas H. Jones
    • 1
  • Stanley C. Winistorfer
    • 1
  1. 1.Department of PediatricsUniversity of IowaIowa City

Personalised recommendations