Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas
Summary: CRISPR-Cas genome engineering in the unicellular green algal model Chlamydomonas reinhardtii has until recently suffered from low integration efficiencies despite traditional genetics being well established. Here, we present a protocol for efficient homology-directed knockin mutagenesis in...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2024-03-01
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| Series: | STAR Protocols |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166723007414 |
| _version_ | 1827583711570296832 |
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| author | Adrian Pascal Nievergelt Dennis Ray Diener Aliona Bogdanova Thomas Brown Gaia Pigino |
| author_facet | Adrian Pascal Nievergelt Dennis Ray Diener Aliona Bogdanova Thomas Brown Gaia Pigino |
| author_sort | Adrian Pascal Nievergelt |
| collection | DOAJ |
| description | Summary: CRISPR-Cas genome engineering in the unicellular green algal model Chlamydomonas reinhardtii has until recently suffered from low integration efficiencies despite traditional genetics being well established. Here, we present a protocol for efficient homology-directed knockin mutagenesis in all commonly used strains of Chlamydomonas. We describe steps for scarless integration of fusion tags and sequence modifications of almost all proteins without the need for a preceding mutant line. We further empower this genetic-editing approach by efficient crossing and highly robust screening protocols.For complete details on the use and execution of this protocol, please refer to Nievergelt et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-03-08T23:10:38Z |
| format | Article |
| id | doaj.art-a20bbe5462d84687855cf795b0406c7d |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-03-08T23:10:38Z |
| publishDate | 2024-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-a20bbe5462d84687855cf795b0406c7d2023-12-15T07:26:03ZengElsevierSTAR Protocols2666-16672024-03-0151102774Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-CasAdrian Pascal Nievergelt0Dennis Ray Diener1Aliona Bogdanova2Thomas Brown3Gaia Pigino4Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, Germany; Human Technopole, V.le Rita Levi-Montalcini, 1, 20017 Milan, Italy; Corresponding authorMax Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, GermanyMax Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, GermanyMax Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, Germany; DRESDEN-concept Genome Center (DcGC), Center for Molecular and Cellular Bioengineering, Technische Universität Dresden, Dresden, GermanyHuman Technopole, V.le Rita Levi-Montalcini, 1, 20017 Milan, Italy; Corresponding authorSummary: CRISPR-Cas genome engineering in the unicellular green algal model Chlamydomonas reinhardtii has until recently suffered from low integration efficiencies despite traditional genetics being well established. Here, we present a protocol for efficient homology-directed knockin mutagenesis in all commonly used strains of Chlamydomonas. We describe steps for scarless integration of fusion tags and sequence modifications of almost all proteins without the need for a preceding mutant line. We further empower this genetic-editing approach by efficient crossing and highly robust screening protocols.For complete details on the use and execution of this protocol, please refer to Nievergelt et al. (2023).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166723007414Cell BiologyCRISPRGeneticsMolecular Biology |
| spellingShingle | Adrian Pascal Nievergelt Dennis Ray Diener Aliona Bogdanova Thomas Brown Gaia Pigino Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas STAR Protocols Cell Biology CRISPR Genetics Molecular Biology |
| title | Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas |
| title_full | Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas |
| title_fullStr | Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas |
| title_full_unstemmed | Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas |
| title_short | Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas |
| title_sort | protocol for precision editing of endogenous chlamydomonas reinhardtii genes with crispr cas |
| topic | Cell Biology CRISPR Genetics Molecular Biology |
| url | http://www.sciencedirect.com/science/article/pii/S2666166723007414 |
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