Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries

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...

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Autores principales:	Emmanouil Metzakopian, Alex Strong, Vivek Iyer, Alex Hodgkins, Konstantinos Tzelepis, Liliana Antunes, Mathias J Friedrich, Qiaohua Kang, Teresa Davidson, Jacob Lamberth, Christina Hoffmann, Gregory D. Davis, George S. Vassiliou, William C. Skarnes, Allan Bradley
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/e8aefb0888d54b0ca7e199e98b82bbba
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spelling	oai:doaj.org-article:e8aefb0888d54b0ca7e199e98b82bbba2021-12-02T12:30:27ZEnhancing the genome editing toolbox: genome wide CRISPR arrayed libraries10.1038/s41598-017-01766-52045-2322https://doaj.org/article/e8aefb0888d54b0ca7e199e98b82bbba2017-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-01766-5https://doaj.org/toc/2045-2322Abstract 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.Emmanouil MetzakopianAlex StrongVivek IyerAlex HodgkinsKonstantinos TzelepisLiliana AntunesMathias J FriedrichQiaohua KangTeresa DavidsonJacob LamberthChristina HoffmannGregory D. DavisGeorge S. VassiliouWilliam C. SkarnesAllan BradleyNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-9 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Emmanouil Metzakopian Alex Strong Vivek Iyer Alex Hodgkins Konstantinos Tzelepis Liliana Antunes Mathias J Friedrich Qiaohua Kang Teresa Davidson Jacob Lamberth Christina Hoffmann Gregory D. Davis George S. Vassiliou William C. Skarnes Allan Bradley Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
description	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.
format	article
author	Emmanouil Metzakopian Alex Strong Vivek Iyer Alex Hodgkins Konstantinos Tzelepis Liliana Antunes Mathias J Friedrich Qiaohua Kang Teresa Davidson Jacob Lamberth Christina Hoffmann Gregory D. Davis George S. Vassiliou William C. Skarnes Allan Bradley
author_facet	Emmanouil Metzakopian Alex Strong Vivek Iyer Alex Hodgkins Konstantinos Tzelepis Liliana Antunes Mathias J Friedrich Qiaohua Kang Teresa Davidson Jacob Lamberth Christina Hoffmann Gregory D. Davis George S. Vassiliou William C. Skarnes Allan Bradley
author_sort	Emmanouil Metzakopian
title	Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
title_short	Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
title_full	Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
title_fullStr	Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
title_full_unstemmed	Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries
title_sort	enhancing the genome editing toolbox: genome wide crispr arrayed libraries
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/e8aefb0888d54b0ca7e199e98b82bbba
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Enhancing the genome editing toolbox: genome wide CRISPR arrayed libraries

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