One-step CRISPR-Cas9 protocol for the generation of plug & play conditional knockouts in Drosophila melanogaster

Joyce J S Yu; Jean-Paul Vincent; Ian J McGough

doi:10.1016/j.xpro.2021.100560

One-step CRISPR-Cas9 protocol for the generation of plug & play conditional knockouts in Drosophila melanogaster

STAR Protoc. 2021 May 25;2(2):100560. doi: 10.1016/j.xpro.2021.100560. eCollection 2021 Jun 18.

Authors

Joyce J S Yu¹, Jean-Paul Vincent¹, Ian J McGough¹

Affiliation

¹ The Francis Crick Institute, London NW1 1AT, UK.

Abstract

This one-step method generates, for any locus, a conditional knockout allele in Drosophila. The allele carries a bright fluorescent marker for easy screening and an attP landing site for subsequent genetic manipulations. After removing the selectable marker with Cre, the attP site can be used to insert DNA fragments expressing tagged or mutant alleles to determine protein localization and function in a tissue- and stage-specific manner. Only a single round of CRISPR-Cas9-mediated editing is required. For complete details on the use and execution of this protocol, please refer to the DWnt4[cKO] example in Yu et al. (2020).

Keywords: CRISPR; Genetics; Model Organisms.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Alleles
Animals
CRISPR-Cas Systems*
Drosophila melanogaster / genetics*
Gene Editing / methods
Gene Knockdown Techniques*

Abstract

Publication types

MeSH terms

Grants and funding