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Updated: Jan 16, 2026
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Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast
Published on: May 15, 2018
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Pichia pastoris Random Mutagenesis and Screening.
Christine Winkler1, Joan Lin-Cereghino2, Geoff P Lin-Cereghino3
1Institute of Molecular Biotechnology, Graz University of Technology, Graz, Austria.
Methods in Molecular Biology (Clifton, N.J.)
|October 1, 2025
Summary
Random mutagenesis in Komagataella phaffii enhances protein secretion. This method identifies novel genes that improve the production of recombinant proteins like horseradish peroxidase and beta-galactosidase.
Area of Science:
- Biotechnology
- Molecular Biology
- Yeast Genetics
Background:
- Random mutagenesis is crucial for understanding gene function and molecular pathways.
- Komagataella phaffii is a methylotrophic yeast utilized for recombinant protein production.
Purpose of the Study:
- To establish protocols for random mutagenesis in Komagataella phaffii.
- To screen for mutants with enhanced recombinant protein secretion.
- To identify novel genes involved in improving protein secretion.
Main Methods:
- Random mutagenesis via non-homologous DNA cassette insertion in K. phaffii.
- Identification of mutagenized loci using cassette-specific primers.
- Screening for enhanced secretion of horseradish peroxidase (HRP) and β-galactosidase.
Main Results:
- Developed and validated two protocols for random mutagenesis in K. phaffii.
- Successfully enhanced the secretion of HRP and β-galactosidase.
- Identified previously unknown genes that positively impact recombinant protein secretion.
Conclusions:
- The developed random mutagenesis system is effective for gene discovery in K. phaffii.
- This methodology holds significant potential for optimizing recombinant protein production in yeast.
- The identified genes offer new targets for further research in protein secretion enhancement.
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