Scalable CRISPR-Cas9 chemical genetic screens in non-transformed human cells

Kevin Lin; Ya-Chu Chang; Ezequiel Marron Fernandez de Velasco; Kevin Wickman; Chad L Myers; Anja-Katrin Bielinsky

doi:10.1016/j.xpro.2022.101675

Scalable CRISPR-Cas9 chemical genetic screens in non-transformed human cells

STAR Protoc. 2022 Dec 16;3(4):101675. doi: 10.1016/j.xpro.2022.101675. Epub 2022 Sep 14.

Authors

Kevin Lin¹, Ya-Chu Chang², Ezequiel Marron Fernandez de Velasco³, Kevin Wickman³, Chad L Myers⁴, Anja-Katrin Bielinsky⁵

Affiliations

¹ Department of Computer Science and Engineering, University of Minnesota, Minneapolis, MN 55455, USA. Electronic address: linx1048@umn.edu.
² Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA. Electronic address: chan1285@umn.edu.
³ Department of Pharmacology, University of Minnesota, Minneapolis, MN 55455, USA.
⁴ Department of Computer Science and Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
⁵ Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA. Electronic address: bieli003@umn.edu.

Abstract

Pooled lentiviral CRISPR-Cas9 screens are utilized for assessing the differential sensitivity or resistance of many single-gene knockouts to a compound. Here, we present a scalable approach for high-throughput compound screening by utilizing a small custom library. We describe steps to perform a proof-of-principle chemical screen in non-transformed hTERT RPE-1 TP53^-/- cells with higher coverage and greater timepoint resolution compared to genome-wide screens. This approach can be adapted for use in various cell lines, compounds, and other focused sgRNA libraries.

Keywords: CRISPR; Genomics; High Throughput Screening; Molecular Biology; Sequencing.

Publication types

Research Support, N.I.H., Extramural
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

CRISPR-Cas Systems*
Gene Knockout Techniques
Gene Library
Genetic Testing*
Genome
Humans

Abstract

Publication types

MeSH terms

Grants and funding