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Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries.

Published version
Peer-reviewed

Type

Article

Change log

Authors

Metzakopian, Emmanouil 
Strong, Alex 
Iyer, Vivek 
Hodgkins, Alex 
Tzelepis, Konstantinos  ORCID logo  https://orcid.org/0000-0002-4865-7648

Abstract

CRISPR-Cas9 technology has accelerated biological research becoming routine for many laboratories. It is rapidly replacing conventional gene editing techniques and has high utility for both genome-wide and gene-focussed applications. Here we present the first individually cloned CRISPR-Cas9 genome wide arrayed sgRNA libraries covering 17,166 human and 20,430 mouse genes at a complexity of 34,332 sgRNAs for human and 40,860 sgRNAs for the mouse genome. For flexibility in generating stable cell lines the sgRNAs have been cloned in a lentivirus backbone containing PiggyBac transposase recognition elements together with fluorescent and drug selection markers. Over 95% of tested sgRNA induced specific DNA cleavage as measured by CEL-1 assays. Furthermore, sgRNA targeting GPI anchor protein pathway genes induced loss of function mutations in human and mouse cell lines measured by FLAER labelling. These arrayed libraries offer the prospect for performing screens on individual genes, combinations as well as larger gene sets. They also facilitate rapid deconvolution of signals from genome-wide screens. This set of vectors provide an organized comprehensive gene editing toolbox of considerable scientific value.

Description

Keywords

Animals, CRISPR-Cas Systems, GPI-Linked Proteins, Gene Editing, Gene Library, Genetic Vectors, Genome-Wide Association Study, Humans, Lentivirus, Mice, Phenotype, RNA, Guide, CRISPR-Cas Systems, Signal Transduction

Journal Title

Sci Rep

Conference Name

Journal ISSN

2045-2322
2045-2322

Volume Title

7

Publisher

Springer Science and Business Media LLC
Sponsorship
Medical Research Council (MC_PC_12009)