Author:
Chu Van Trung,Graf Robin,Wirtz Tristan,Weber Timm,Favret Jeremy,Li Xun,Petsch Kerstin,Tran Ngoc Tung,Sieweke Michael H.,Berek Claudia,Kühn Ralf,Rajewsky Klaus
Abstract
Applying clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)-mediated mutagenesis to primary mouse immune cells, we used high-fidelity single guide RNAs (sgRNAs) designed with an sgRNA design tool (CrispRGold) to target genes in primary B cells, T cells, and macrophages isolated from a Cas9 transgenic mouse line. Using this system, we achieved an average knockout efficiency of 80% in B cells. On this basis, we established a robust small-scale CRISPR-mediated screen in these cells and identified genes essential for B-cell activation and plasma cell differentiation. This screening system does not require deep sequencing and may serve as a precedent for the application of CRISPR/Cas9 to primary mouse cells.
Funder
EC | European Research Council
Bundesministerium für Forschung und Technologie
Agence Nationale de la Recherche
Publisher
Proceedings of the National Academy of Sciences
Cited by
106 articles.
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