High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.

Gene editing in C. elegans using plasmid-based CRISPR reagents requires microinjection of many animals to produce a single edit. Germline silencing of plasmid-borne Cas9 is a major cause of inefficient editing. Here, we present a set of C. elegans strains that constitutively express Cas9 in the germ...

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Autores principales:	Matthew L Schwartz, M Wayne Davis, Matthew S Rich, Erik M Jorgensen
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Genetics QH426-470
Acceso en línea:	https://doaj.org/article/777ecedabc114ade999538b172ce2a3a
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id	oai:doaj.org-article:777ecedabc114ade999538b172ce2a3a
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spelling	oai:doaj.org-article:777ecedabc114ade999538b172ce2a3a2021-12-02T20:03:16ZHigh-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.1553-73901553-740410.1371/journal.pgen.1009755https://doaj.org/article/777ecedabc114ade999538b172ce2a3a2021-11-01T00:00:00Zhttps://doi.org/10.1371/journal.pgen.1009755https://doaj.org/toc/1553-7390https://doaj.org/toc/1553-7404Gene editing in C. elegans using plasmid-based CRISPR reagents requires microinjection of many animals to produce a single edit. Germline silencing of plasmid-borne Cas9 is a major cause of inefficient editing. Here, we present a set of C. elegans strains that constitutively express Cas9 in the germline from an integrated transgene. These strains markedly improve the success rate for plasmid-based CRISPR edits. For simple, short homology arm GFP insertions, 50-100% of injected animals typically produce edited progeny, depending on the target locus. Template-guided editing from an extrachromosomal array is maintained over multiple generations. We have built strains with the Cas9 transgene on multiple chromosomes. Additionally, each Cas9 locus also contains a heatshock-driven Cre recombinase for selectable marker removal and a bright fluorescence marker for easy outcrossing. These integrated Cas9 strains greatly reduce the workload for producing individual genome edits.Matthew L SchwartzM Wayne DavisMatthew S RichErik M JorgensenPublic Library of Science (PLoS)articleGeneticsQH426-470ENPLoS Genetics, Vol 17, Iss 11, p e1009755 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Genetics QH426-470
spellingShingle	Genetics QH426-470 Matthew L Schwartz M Wayne Davis Matthew S Rich Erik M Jorgensen High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
description	Gene editing in C. elegans using plasmid-based CRISPR reagents requires microinjection of many animals to produce a single edit. Germline silencing of plasmid-borne Cas9 is a major cause of inefficient editing. Here, we present a set of C. elegans strains that constitutively express Cas9 in the germline from an integrated transgene. These strains markedly improve the success rate for plasmid-based CRISPR edits. For simple, short homology arm GFP insertions, 50-100% of injected animals typically produce edited progeny, depending on the target locus. Template-guided editing from an extrachromosomal array is maintained over multiple generations. We have built strains with the Cas9 transgene on multiple chromosomes. Additionally, each Cas9 locus also contains a heatshock-driven Cre recombinase for selectable marker removal and a bright fluorescence marker for easy outcrossing. These integrated Cas9 strains greatly reduce the workload for producing individual genome edits.
format	article
author	Matthew L Schwartz M Wayne Davis Matthew S Rich Erik M Jorgensen
author_facet	Matthew L Schwartz M Wayne Davis Matthew S Rich Erik M Jorgensen
author_sort	Matthew L Schwartz
title	High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
title_short	High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
title_full	High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
title_fullStr	High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
title_full_unstemmed	High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.
title_sort	high-efficiency crispr gene editing in c. elegans using cas9 integrated into the genome.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/777ecedabc114ade999538b172ce2a3a
work_keys_str_mv	AT matthewlschwartz highefficiencycrisprgeneeditingincelegansusingcas9integratedintothegenome AT mwaynedavis highefficiencycrisprgeneeditingincelegansusingcas9integratedintothegenome AT matthewsrich highefficiencycrisprgeneeditingincelegansusingcas9integratedintothegenome AT erikmjorgensen highefficiencycrisprgeneeditingincelegansusingcas9integratedintothegenome
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High-efficiency CRISPR gene editing in C. elegans using Cas9 integrated into the genome.

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