ArticleWorld journal of microbiology & biotechnology2019
Construction of a series of episomal plasmids and their application in the development of an efficient CRISPR/Cas9 system in Pichia pastoris.
Article in World journal of microbiology & biotechnology, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 27 papers.
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Who cites it
27 citing papers in PubMed, 53 citations in OpenAlex.
- Development and characterization of a Cre/Synthetic and systems biotechnology · 2027Article
- Expanding the Genetic Toolkit of Pichia kudriavzevii E1 for Enhanced Mycoprotein Production.Microbial biotechnology · 2026Article
- Metabolic engineering ofApplied and environmental microbiology · 2026Review
- CRISPR-Cas9-based one-step multiplexed genome editing through optimizing guide RNA processing strategies inSynthetic and systems biotechnology · 2025Article
- Advancing recombinant protein expression in Komagataella phaffii: opportunities and challenges.FEMS yeast research · 2025Review
- CRISPR-Cas9 Gene Therapy: Non-Viral Delivery and Stimuli-Responsive Nanoformulations.Molecules (Basel, Switzerland) · 2025Review
- Screening and characterization of integration sites based on CRISPR-Cpf1 inSynthetic and systems biotechnology · 2024Article
- Cytokinetic engineering enhances the secretory production of recombinant human lysozyme in Komagataella phaffii.Microbial cell factories · 2024Article
- Oligonucleotide-based CRISPR-Cas9 toolbox for efficient engineering of Komagataella phaffii.FEMS yeast research · 2024Article
- Multiplex Marker-Less Genome Integration in Pichia pastoris Using CRISPR/Cas9.Methods in molecular biology (Clifton, N.J.) · 2024Article
- Current achievements, strategies, obstacles, and overcoming the challenges of the protein engineering in Pichia pastoris expression system.World journal of microbiology & biotechnology · 2023Review
- Genetic tools for metabolic engineering ofEngineering microbiology · 2023Review
- Advances in Metabolic Engineering ofJournal of fungi (Basel, Switzerland) · 2023Review
- CRISPR-Cas Technology for Bioengineering Conventional and Non-Conventional Yeasts: Progress and New Challenges.International journal of molecular sciences · 2023Review
- A novel CRISPR/Cas9 system with high genomic editing efficiency and recyclable auxotrophic selective marker for multiple-step metabolic rewriting inSynthetic and systems biotechnology · 2023Article
- Synthetic methylotrophic yeasts for the sustainable fuel and chemical production.Biotechnology for biofuels and bioproducts · 2022Review
- Comparative transcriptome and metabolome analyses reveal the methanol dissimilation pathway of Pichia pastoris.BMC genomics · 2022Article
- Current advances ofFrontiers in microbiology · 2022Review
- Recent Molecular Tools for the Genetic Manipulation of Highly Industrially Important Mucoromycota Fungi.Journal of fungi (Basel, Switzerland) · 2021Review
- Development of synthetic biology tools to engineerSynthetic and systems biotechnology · 2021Review
Corrections and comments
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Authors and funding
8 authors at 3 institutions in 3 countries.
Funding
Abstract
The methylotrophic yeast Pichia pastoris is widely used in recombinant expression of eukaryotic proteins owing to the ability of post-translational modification, tightly regulated promoters, and high cell density fermentation. However, episomal plasmids for heterologous gene expression and the CRISPR/Cas9 system for genome editing have not been well developed in P. pastoris. In the present study, a panel of episomal plasmids containing various autonomously replicating sequences (ARSs) were constructed and their performance in transformation efficiency, copy numbers, and propagation stability were systematically compared. Among the five ARSs with different origins, panARS isolated from Kluyveromyces lactis was determined to have the best performance and used to develop an efficient CRISPR/Cas9 based genome editing system. Compared with a previously reported system using the endogenous and most commonly used ARS (PARS1), the CRISPR/Cas9 genome editing efficiency was increased for more than tenfold. Owing to the higher plasmid stability with panARS, efficient CRISPR/Cas9-mediated genome editing with a type III promoter (i.e. SER promoter) to drive the expression of the single guide RNA (sgRNA) was achieved for the first time. The constructed episomal plasmids and developed CRISPR/Cas9 system will be important synthetic biology tools for both fundamental studies and industrial applications of P. pastoris.
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Registered trials
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