Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas.
| Autor: | Nievergelt AP; Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, Germany; Human Technopole, V.le Rita Levi-Montalcini, 1, 20017 Milan, Italy. Electronic address: adrian@nievergelt-merz.ch., Diener DR; Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, Germany., Bogdanova A; Max Planck Institute of Molecular Cell Biology and Genetics, Pfotenhauerstraße 108, 01307 Dresden, Germany., Brown T; Max 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, Germany., Pigino G; Human Technopole, V.le Rita Levi-Montalcini, 1, 20017 Milan, Italy. Electronic address: gaia.pigino@fht.org. |
|---|---|
| Jazyk: | angličtina |
| Zdroj: | STAR protocols [STAR Protoc] 2024 Mar 15; Vol. 5 (1), pp. 102774. Date of Electronic Publication: 2023 Dec 13. |
| DOI: | 10.1016/j.xpro.2023.102774 |
| Abstrakt: | 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 . Competing Interests: Declaration of interests The authors declare no competing interests. (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.) |
| Databáze: | MEDLINE |
| Externí odkaz: |
|