Expression in the yeast Pichia pastoris.
The yeast Pichia pastoris has become the premier example of yeast species used for the production of recombinant proteins. Advantages of this yeast for expression include tightly regulated and efficient promoters and a strong tendency for respiratory growth as opposed to fermentative growth. This chapter assumes the reader is proficient in molecular biology and details the more yeast specific procedures involved in utilizing the P. pastoris system for gene expression. Procedures to be found here include: strain construction by classical yeast genetics, the logic in selection of a vector and strain, preparation of electrocompetent yeast cells and transformation by electroporation, and the yeast colony western blot or Yeastern blot method for visualizing secreted proteins around yeast colonies.
Cregg
James M
JM
Keck Graduate Institute of Applied Life Sciences, Claremont, California, USA.
Tolstorukov
Ilya
I
Kusari
Anasua
A
Sunga
Jay
J
Madden
Knut
K
Chappell
Thomas
T
D000818:Animals; D003001:Cloning, Molecular / Q000379:methods; D015966:Gene Expression Regulation, Fungal; D018014:Gene Transfer Techniques; D006801:Humans; D008828:Microbiological Techniques / Q000379:methods*; D008957:Models, Genetic; D010843:Pichia / Q000737:chemistry / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000502:physiology; D011994:Recombinant Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D011994:Recombinant Proteins