Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.

Improved genome engineering methods that enable automation of large and precise edits are essential for systematic investigations of genome function. We adapted peel-1 negative selection to an optimized Dual-Marker Selection (DMS) cassette protocol for CRISPR-Cas9 genome engineering in Caenorhabditi...

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Autores principales:	Troy A McDiarmid, Vinci Au, Donald G Moerman, Catharine H Rankin
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f0cebc28659b427cbe8025590b05f26a
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spelling	oai:doaj.org-article:f0cebc28659b427cbe8025590b05f26a2021-12-02T20:15:51ZPeel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.1932-620310.1371/journal.pone.0238950https://doaj.org/article/f0cebc28659b427cbe8025590b05f26a2020-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0238950https://doaj.org/toc/1932-6203Improved genome engineering methods that enable automation of large and precise edits are essential for systematic investigations of genome function. We adapted peel-1 negative selection to an optimized Dual-Marker Selection (DMS) cassette protocol for CRISPR-Cas9 genome engineering in Caenorhabditis elegans and observed robust increases in multiple measures of efficiency that were consistent across injectors and four genomic loci. The use of Peel-1-DMS selection killed animals harboring transgenes as extrachromosomal arrays and spared genome-edited integrants, often circumventing the need for visual screening to identify genome-edited animals. To demonstrate the applicability of the approach, we created deletion alleles in the putative proteasomal subunit pbs-1 and the uncharacterized gene K04F10.3 and used machine vision to automatically characterize their phenotypic profiles, revealing homozygous essential and heterozygous behavioral phenotypes. These results provide a robust and scalable approach to rapidly generate and phenotype genome-edited animals without the need for screening or scoring by eye.Troy A McDiarmidVinci AuDonald G MoermanCatharine H RankinPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 15, Iss 9, p e0238950 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Troy A McDiarmid Vinci Au Donald G Moerman Catharine H Rankin Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
description	Improved genome engineering methods that enable automation of large and precise edits are essential for systematic investigations of genome function. We adapted peel-1 negative selection to an optimized Dual-Marker Selection (DMS) cassette protocol for CRISPR-Cas9 genome engineering in Caenorhabditis elegans and observed robust increases in multiple measures of efficiency that were consistent across injectors and four genomic loci. The use of Peel-1-DMS selection killed animals harboring transgenes as extrachromosomal arrays and spared genome-edited integrants, often circumventing the need for visual screening to identify genome-edited animals. To demonstrate the applicability of the approach, we created deletion alleles in the putative proteasomal subunit pbs-1 and the uncharacterized gene K04F10.3 and used machine vision to automatically characterize their phenotypic profiles, revealing homozygous essential and heterozygous behavioral phenotypes. These results provide a robust and scalable approach to rapidly generate and phenotype genome-edited animals without the need for screening or scoring by eye.
format	article
author	Troy A McDiarmid Vinci Au Donald G Moerman Catharine H Rankin
author_facet	Troy A McDiarmid Vinci Au Donald G Moerman Catharine H Rankin
author_sort	Troy A McDiarmid
title	Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
title_short	Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
title_full	Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
title_fullStr	Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
title_full_unstemmed	Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.
title_sort	peel-1 negative selection promotes screening-free crispr-cas9 genome editing in caenorhabditis elegans.
publisher	Public Library of Science (PLoS)
publishDate	2020
url	https://doaj.org/article/f0cebc28659b427cbe8025590b05f26a
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Peel-1 negative selection promotes screening-free CRISPR-Cas9 genome editing in Caenorhabditis elegans.

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