Protocol for in vivo CRISPR screening using selective CRISPR antigen removal lentiviral vectors

Sarah Kate Lane-Reticker; Emily A Kessler; Audrey J Muscato; Sarah Y Kim; John G Doench; Kathleen B Yates; Robert T Manguso; Juan Dubrot

doi:10.1016/j.xpro.2023.102082

Protocol for in vivo CRISPR screening using selective CRISPR antigen removal lentiviral vectors

STAR Protoc. 2023 Mar 17;4(1):102082. doi: 10.1016/j.xpro.2023.102082. Epub 2023 Feb 1.

Authors

Sarah Kate Lane-Reticker¹, Emily A Kessler¹, Audrey J Muscato¹, Sarah Y Kim¹, John G Doench¹, Kathleen B Yates², Robert T Manguso³, Juan Dubrot⁴

Affiliations

¹ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
² Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Center for Cancer Research, Massachusetts General Hospital, Boston, MA 02114, USA.
³ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Center for Cancer Research, Massachusetts General Hospital, Boston, MA 02114, USA. Electronic address: rmanguso@broadinstitute.org.
⁴ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; Solid Tumors Program, Division of Oncology, Center for Applied Medical Research (CIMA), University of Navarra, Pamplona, Spain. Electronic address: jdubrot@unav.es.

Abstract

Recognition of Cas9 and other proteins encoded in delivery vectors has limited CRISPR technology in vivo. Here, we present a protocol for genome engineering using selective CRISPR antigen removal (SCAR) lentiviral vectors in Renca mouse model. This protocol describes how to conduct an in vivo genetic screen with a sgRNA library and SCAR vectors that can be applied to different cell lines and contexts. For complete details on the use and execution of this protocol, please refer to Dubrot et al. (2021).¹.

Keywords: CRISPR; Cancer; Immunology; Sequencing.

Publication types

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

MeSH terms

Animals
CRISPR-Cas Systems* / genetics
Cell Line
Gene Library
Genome
Mice
RNA, Guide, CRISPR-Cas Systems*

Substances

RNA, Guide, CRISPR-Cas Systems