Pichia pastoris as a host system for transformations
| Autor: | A Y Hessler, Kevin J. Barringer, James M. Cregg, Knut Madden |
|---|---|
| Rok vydání: | 1985 |
| Předmět: |
biology
Base Sequence Autonomously replicating sequence Mutant Genes Fungal Fungal genetics Chromosome Mapping Saccharomyces cerevisiae Cell Biology biology.organism_classification Histidinol dehydrogenase Molecular biology Pichia Pichia pastoris Transformation (genetics) Alcohol Oxidoreductases Plasmid Transformation Genetic Mutation Saccharomycetales DNA Fungal Molecular Biology Selectable marker Research Article Plasmids |
| Zdroj: | Molecular and Cellular Biology. 5:3376-3385 |
| ISSN: | 1098-5549 0270-7306 |
| DOI: | 10.1128/mcb.5.12.3376 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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