Protocol for precision editing of endogenous Chlamydomonas reinhardtii genes with CRISPR-Cas

STAR Protoc. 2024 Mar 15;5(1):102774. doi: 10.1016/j.xpro.2023.102774. Epub 2023 Dec 13.


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.

Keywords: CRISPR; Cell Biology; Genetics; Molecular Biology.

MeSH terms

  • CRISPR-Cas Systems* / genetics
  • Chlamydomonas reinhardtii* / genetics
  • Chlamydomonas reinhardtii* / metabolism
  • Gene Editing / methods
  • Genome
  • Mutagenesis