Microinjection for precision genome editing in Caenorhabditis elegans

Krishna S Ghanta; Takao Ishidate; Craig C Mello

doi:10.1016/j.xpro.2021.100748

Microinjection for precision genome editing in Caenorhabditis elegans

STAR Protoc. 2021 Sep 1;2(3):100748. doi: 10.1016/j.xpro.2021.100748. eCollection 2021 Sep 17.

Authors

Krishna S Ghanta¹, Takao Ishidate^{1

2}, Craig C Mello^{1

3

2}

Affiliations

¹ RNA Therapeutics Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
² Howard Hughes Medical Institute, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.
³ Program in Molecular Medicine, University of Massachusetts Medical School, 368 Plantation Street, Worcester, MA 01605, USA.

Abstract

In Caenorhabditis elegans, targeted genome editing techniques are now routinely used to generate germline edits. The remarkable ease of C. elegans germline editing is attributed to the syncytial nature of the pachytene ovary which is easily accessed by microinjection. This protocol describes the step-by-step details and troubleshooting tips for the entire CRISPR-Cas genome editing procedure, including gRNA design and microinjection of ribonucleoprotein complexes, followed by screening and genotyping in C. elegans, to help accessing this powerful genetic animal system. For complete details on the use and execution of this protocol, please refer to Ghanta and Mello (2020).

Keywords: CRISPR; genetics; model organisms; molecular biology.

Publication types

Research Support, N.I.H., Extramural

MeSH terms

Animals
CRISPR-Associated Protein 9 / genetics
CRISPR-Cas Systems
Caenorhabditis elegans / genetics
Caenorhabditis elegans Proteins / genetics
Gene Editing / methods*
Genetic Engineering / methods*
Microinjections / methods*

Substances

Caenorhabditis elegans Proteins
CRISPR-Associated Protein 9

Abstract

Publication types

MeSH terms

Substances

Grants and funding