Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages

Abstract Genome engineering using CRISPR/Cas9 technology enables simple, efficient and precise genomic modifications in human cells. Conventional immortalized cell lines can be easily edited or screened using genome-wide libraries with lentiviral transduction. However, cell types derived from the di...

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Autores principales:	Elena Navarro-Guerrero, Chwen Tay, Justin P. Whalley, Sally A. Cowley, Ben Davies, Julian C. Knight, Daniel Ebner
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/d5d3e85be29a477a8b861f701127b798
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spelling	oai:doaj.org-article:d5d3e85be29a477a8b861f701127b7982021-12-02T14:03:45ZGenome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages10.1038/s41598-021-82137-z2045-2322https://doaj.org/article/d5d3e85be29a477a8b861f701127b7982021-02-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-82137-zhttps://doaj.org/toc/2045-2322Abstract Genome engineering using CRISPR/Cas9 technology enables simple, efficient and precise genomic modifications in human cells. Conventional immortalized cell lines can be easily edited or screened using genome-wide libraries with lentiviral transduction. However, cell types derived from the differentiation of induced Pluripotent Stem Cells (iPSC), which often represent more relevant, patient-derived models for human pathology, are much more difficult to engineer as CRISPR/Cas9 delivery to these differentiated cells can be inefficient and toxic. Here, we present an efficient, lentiviral transduction protocol for delivery of CRISPR/Cas9 to macrophages derived from human iPSC with efficiencies close to 100%. We demonstrate CRISPR/Cas9 knockouts for three nonessential proof-of-concept genes—HPRT1, PPIB and CDK4. We then scale the protocol and validate for a genome-wide pooled CRISPR/Cas9 loss-of-function screen. This methodology enables, for the first time, systematic exploration of macrophage involvement in immune responses, chronic inflammation, neurodegenerative diseases and cancer progression, using efficient genome editing techniques.Elena Navarro-GuerreroChwen TayJustin P. WhalleySally A. CowleyBen DaviesJulian C. KnightDaniel EbnerNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-11 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Elena Navarro-Guerrero Chwen Tay Justin P. Whalley Sally A. Cowley Ben Davies Julian C. Knight Daniel Ebner Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
description	Abstract Genome engineering using CRISPR/Cas9 technology enables simple, efficient and precise genomic modifications in human cells. Conventional immortalized cell lines can be easily edited or screened using genome-wide libraries with lentiviral transduction. However, cell types derived from the differentiation of induced Pluripotent Stem Cells (iPSC), which often represent more relevant, patient-derived models for human pathology, are much more difficult to engineer as CRISPR/Cas9 delivery to these differentiated cells can be inefficient and toxic. Here, we present an efficient, lentiviral transduction protocol for delivery of CRISPR/Cas9 to macrophages derived from human iPSC with efficiencies close to 100%. We demonstrate CRISPR/Cas9 knockouts for three nonessential proof-of-concept genes—HPRT1, PPIB and CDK4. We then scale the protocol and validate for a genome-wide pooled CRISPR/Cas9 loss-of-function screen. This methodology enables, for the first time, systematic exploration of macrophage involvement in immune responses, chronic inflammation, neurodegenerative diseases and cancer progression, using efficient genome editing techniques.
format	article
author	Elena Navarro-Guerrero Chwen Tay Justin P. Whalley Sally A. Cowley Ben Davies Julian C. Knight Daniel Ebner
author_facet	Elena Navarro-Guerrero Chwen Tay Justin P. Whalley Sally A. Cowley Ben Davies Julian C. Knight Daniel Ebner
author_sort	Elena Navarro-Guerrero
title	Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
title_short	Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
title_full	Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
title_fullStr	Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
title_full_unstemmed	Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages
title_sort	genome-wide crispr/cas9-knockout in human induced pluripotent stem cell (ipsc)-derived macrophages
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/d5d3e85be29a477a8b861f701127b798
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Genome-wide CRISPR/Cas9-knockout in human induced Pluripotent Stem Cell (iPSC)-derived macrophages

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