Engineering Pichia pastoris Strains Using CRISPR/Cas9 Technologies: The Basic Protocol
Jazyk angličtina Země Spojené státy americké Médium print
Typ dokumentu časopisecké články
- Klíčová slova
- CRISPR/Cas9, Gene knockout, Genome editing, Genome engineering, Guide RNA, Homologous recombination, Komagataella phaffii, Pichia pastoris, Synthetic biology,
- MeSH
- CRISPR-Cas systémy * genetika MeSH
- editace genu * metody MeSH
- genom fungální MeSH
- metabolické inženýrství * metody MeSH
- Pichia * genetika MeSH
- Saccharomycetales * genetika MeSH
- vodící RNA, systémy CRISPR-Cas genetika MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- vodící RNA, systémy CRISPR-Cas MeSH
The CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats, CRISPR-associated protein 9) system has become a commonly used tool for genome editing and metabolic engineering. For Komagataella phaffii, commercialized as Pichia pastoris, the CRISPR/Cas9 protocol for genome editing was established in 2016 and since then has been employed to facilitate genetic modifications such as markerless gene disruptions and deletions as well as to enhance the efficiency of homologous recombination.In this chapter, we describe a robust basic protocol for CRISPR-based genome editing, demonstrating near 100% targeting efficiency for gene inactivation via a frameshift mutation. As described in other chapters of this volume, CRISPR/Cas9 technologies for use in P. pastoris have been further optimized for various specific purposes.
Graz University of Technology Institute of Molecular Biotechnology Graz Austria
University of Chemistry and Technology Prague Department of Biotechnology Prague Czech Republic
Zobrazit více v PubMed
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