Work

Exploiting CRISPR/Cas9 to engineer precise segmental deletions in mouse embryonic stem cells

Year: 2022

Type: article

Abstract: In this protocol, we use CRISPR/Cas9 to generate large deletions of the entire coding region of a gene of interest, generating a hemizygous cell line. Next, we systematically engineer precise in-frame deletions within the intact wild-type allele, facilitating study of multi-domain proteins. The optimized protocol described here allows us to rapidly screen for effective sgRNA pairs and to engineer either an in-frame deletion or a frameshift mutation in high frequencies in mouse embryonic stem cells. For complete details on the use and execution of this protocol, please refer to Panday et al. (2021).

Source: STAR Protocols

DOI: 10.1016/j.xpro.2022.101551

Authors: Rajula Elango, Arvind Panday, Nicholas A. Willis, Ralph Scully

Institutions: Beth Israel Deaconess Medical Center, Harvard University

Language: English

Topic: CRISPR and Genetic Engineering

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: gold

Estimated APC paid: $1,500

Funders: American Association for Cancer Research, National Institutes of Health, National Cancer Institute, National Institute of General Medical Sciences