Pooled CRISPR-activation screening coupled with single-cell RNA-seq in mouse embryonic stem cells

Celia Alda-Catalinas; Melanie A Eckersley-Maslin; Wolf Reik

doi:10.1016/j.xpro.2021.100426

Pooled CRISPR-activation screening coupled with single-cell RNA-seq in mouse embryonic stem cells

STAR Protoc. 2021 Apr 9;2(2):100426. doi: 10.1016/j.xpro.2021.100426. eCollection 2021 Jun 18.

Authors

Celia Alda-Catalinas¹, Melanie A Eckersley-Maslin¹, Wolf Reik^{1

2

3}

Affiliations

¹ Epigenetics Programme, Babraham Institute, Cambridge CB22 3AT, UK.
² Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK.
³ Centre for Trophoblast Research, University of Cambridge, Cambridge CB2 3EG, UK.

Abstract

CRISPR/Cas9 screens are a powerful approach to identify key regulators of biological processes. By combining pooled CRISPR/Cas9 screening with single-cell RNA-sequencing readout, individual perturbations can be assessed in parallel both comprehensively and at scale. Importantly, this allows gene function and regulation to be interrogated at a cellular level in an unbiased manner. Here, we present a protocol to perform pooled CRISPR-activation screens in mouse embryonic stem cells using 10× Genomics scRNA-seq as a readout. For complete information on the generation and use of this protocol, please refer to Alda-Catalinas et al. (2020).

Keywords: CRISPR; Cell culture; Gene Expression; High Throughput Screening; RNAseq; Sequencing; Single Cell; Stem Cells.

Publication types

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

MeSH terms

Animals
CRISPR-Cas Systems / genetics*
Cells, Cultured
Genomics
Mice
Mouse Embryonic Stem Cells / cytology*
Mouse Embryonic Stem Cells / metabolism
RNA-Seq / methods*
Single-Cell Analysis / methods*
Transcriptome / genetics*

Abstract

Publication types

MeSH terms

Grants and funding