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Electroporation of Pichia pastoris

  • Knut Madden
  • Ilya Tolstorukov
  • James Cregg
Chapter
Part of the Fungal Biology book series (FUNGBIO)

Abstract

The introduction of foreign DNA into the yeast Pichia pastoris is similar to the processes used with Saccharomyces cerevisiae and routinely performed by the electroporation of plasmid DNA into electro-competent (E-comp) P. pastoris cells (Becker and Guarente, Methods Enzymol 194:182–187, 1991). Transformation of spheroplasts is more difficult (Cregg et al., Mol Cell Biol 5:3376–3385, 1985), and current methods to make chemically competent P. pastoris cells results in significantly fewer transformants (Pichia protocols, 2nd edn, Totowa, 27–42). Following are protocols to prepare E-comp P. pastoris cells, and to electroporate plasmid DNA into these cells to generate transformed cells.

Keywords

Pichia pastoris Transformation Electroporation Competent cells Spheroplasts 

References

  1. Becker DM, Guarente L (1991) High-efficiency transformation of yeast by electroporation. Methods Enzymol 194:182–187PubMedCrossRefGoogle Scholar
  2. Cregg J (2007) DNA-mediated transformation. In: Cregg J (ed) Pichia protocols, 2nd edn. Humana Press Inc., Totowa, pp 27–42Google Scholar
  3. Cregg JM, Barringer KJ, Hessler AY, Madden KR (1985) Pichia pastoris as a host system for transformations. Mol Cell Biol 5:3376–3385PubMedPubMedCentralGoogle Scholar
  4. Cregg JM, Tolstorukov II, Kusari A, Sunga J, Madden K, Chappell T (2009) Expression in the yeast Pichia pastoris. Methods Enzymol 463:169–189PubMedCrossRefGoogle Scholar
  5. Lin-Cereghino J, Lin-Cereghino GP (2007) Vectors and strains for expression. In: Cregg J (ed) Pichia protocols, 2nd edn. Humana Press Inc., Totowa, pp 11–25Google Scholar

Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  • Knut Madden
    • 1
  • Ilya Tolstorukov
    • 1
    • 2
  • James Cregg
    • 1
    • 2
  1. 1.BioGrammatics, Inc.CarlsbadUSA
  2. 2.Keck Graduate InstituteClaremontUSA

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