title: Design, execution, and analysis of pooled in vitro CRISPR/Cas9 screens.
abstract: The recently described clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology has proven to be an exquisitely powerful and invaluable method of genetic manipulation and/or modification. As such, many researchers have realized the potential of using the CRISPR/Cas9 system as a novel screening method for the identification of important proteins in biological processes and have designed short guide RNA libraries for an in vitro screening. The seminal papers describing these libraries offer valuable information regarding methods for generating the short guide RNA libraries, creating cell lines containing these libraries, and specific details regarding the screening workflow. However, certain considerations are often overlooked that may be important when planning and performing a screen, including which CRISPR library to use and how to best analyze the resulting screen data. In this review, we offer suggestions to answer some of these questions that are not covered as deeply in the papers describing the available CRISPR libraries for an in vitro screening.
authors.affiliation: Pharmacology Graduate Training Program, Department of Pharmacology and Molecular Sciences, Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Linde A
authors.initials: LA
authors.lastname: Miles
authors.affiliation: RNAi Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY, USA.
authors.forename: Ralph J
authors.initials: RJ
authors.lastname: Garippa
authors.affiliation: Department of Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA.|Molecular Pharmacology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA.|Weill Cornell Medical College, New York, NY, USA.
authors.forename: John T
authors.initials: JT
authors.lastname: Poirier
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems; D002460:Cell Line; D000072669:Gene Editing; D055786:Gene Knockout Techniques; D015723:Gene Library; D005820:Genetic Testing; D006801:Humans; D034622:RNA Interference; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics; D015533:Transcriptional Activation
chemical_list: D000094704:RNA, Guide, CRISPR-Cas Systems
keywords: CRISPR; Libraries; Screening; sgRNA