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PMID: 3915774 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Pichia pastoris as a host system for transformations.

Molecular and cellular biology ·Vol. 5 ·No. 12 ·1985-12-00 ·Pages 3376-85

Cregg JM, Barringer KJ, Hessler AY, Madden KR

Abstract

We developed a methylotrophic yeast, Pichia pastoris, as a host for DNA transformations. The system is based on an auxotrophic mutant host of P. pastoris which is defective in histidinol dehydrogenase. As a selectable marker, we isolated and characterized the P. pastoris HIS4 gene. Plasmid vectors which contained either the P. pastoris or the Saccharomyces cerevisiae HIS4 gene transformed the P. pastoris mutant host. DNA transfer was accomplished by a modified version of the spheroplast generation (CaCl2-polyethylene glycol)-fusion procedure developed for S. cerevisiae. In addition, we report the isolation and characterization of P. pastoris DNA fragments with autonomous replication sequence activity. Two fragments, PARS1 and PARS2, when present on plasmids increased transformation frequencies to 10(5)/micrograms and maintained the plasmids as autonomous elements in P. pastoris cells.

MeSH Terms
Alcohol Oxidoreductases/genetics Base Sequence Chromosome Mapping DNA, Fungal/genetics Genes, Fungal Mutation Pichia/genetics Plasmids Saccharomyces cerevisiae/genetics Saccharomycetales/genetics Transformation, Genetic
Chemicals
DNA, Fungal Alcohol Oxidoreductases histidinol dehydrogenase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Cregg J M
Barringer K J
Hessler A Y
Madden K R
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34 references, click to expand
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.5.12.3376-3385.1985
Published
1985-12-00
Pages
3376-85
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC369166
Subset
IM
Databases
GENBANK
M11199, M11201
External Links
PubMed source DOI: 10.1128/mcb.5.12.3376-3385.1985 PMC Full Text View journal details
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