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Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops

Genome editing has become a major tool for both functional studies and plant breeding in several species. Besides generating knockouts through the classical CRISPR-Cas9 system, recent development of CRISPR base editing holds great and exciting opportunities for the production of gain-of-function mut...

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Autores principales: Veillet, Florian, Perrot, Laura, Guyon-Debast, Anouchka, Kermarrec, Marie-Paule, Chauvin, Laura, Chauvin, Jean-Eric, Gallois, Jean-Luc, Mazier, Marianne, Nogué, Fabien
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7036883/
https://www.ncbi.nlm.nih.gov/pubmed/32033083
http://dx.doi.org/10.3390/ijms21031024
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author Veillet, Florian
Perrot, Laura
Guyon-Debast, Anouchka
Kermarrec, Marie-Paule
Chauvin, Laura
Chauvin, Jean-Eric
Gallois, Jean-Luc
Mazier, Marianne
Nogué, Fabien
author_facet Veillet, Florian
Perrot, Laura
Guyon-Debast, Anouchka
Kermarrec, Marie-Paule
Chauvin, Laura
Chauvin, Jean-Eric
Gallois, Jean-Luc
Mazier, Marianne
Nogué, Fabien
author_sort Veillet, Florian
collection PubMed
description Genome editing has become a major tool for both functional studies and plant breeding in several species. Besides generating knockouts through the classical CRISPR-Cas9 system, recent development of CRISPR base editing holds great and exciting opportunities for the production of gain-of-function mutants. The PAM requirement is a strong limitation for CRISPR technologies such as base editing, because the base substitution mainly occurs in a small edition window. As precise single amino-acid substitution can be responsible for functions associated to some domains or agronomic traits, development of Cas9 variants with relaxed PAM recognition is of upmost importance for gene function analysis and plant breeding. Recently, the SpCas9-NG variant that recognizes the NGN PAM has been successfully tested in plants, mainly in monocotyledon species. In this work, we studied the efficiency of SpCas9-NG in the model moss Physcomitrella patens and two Solanaceae crops (Solanum lycopersicum and Solanum tuberosum) for both classical CRISPR-generated gene knock-out and cytosine base editing. We showed that the SpCas9-NG greatly expands the scope of genome editing by allowing the targeting of non-canonical NGT and NGA PAMs. The CRISPR toolbox developed in our study opens up new gene function analysis and plant breeding perspectives for model and crop plants.
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spelling pubmed-70368832020-03-11 Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops Veillet, Florian Perrot, Laura Guyon-Debast, Anouchka Kermarrec, Marie-Paule Chauvin, Laura Chauvin, Jean-Eric Gallois, Jean-Luc Mazier, Marianne Nogué, Fabien Int J Mol Sci Communication Genome editing has become a major tool for both functional studies and plant breeding in several species. Besides generating knockouts through the classical CRISPR-Cas9 system, recent development of CRISPR base editing holds great and exciting opportunities for the production of gain-of-function mutants. The PAM requirement is a strong limitation for CRISPR technologies such as base editing, because the base substitution mainly occurs in a small edition window. As precise single amino-acid substitution can be responsible for functions associated to some domains or agronomic traits, development of Cas9 variants with relaxed PAM recognition is of upmost importance for gene function analysis and plant breeding. Recently, the SpCas9-NG variant that recognizes the NGN PAM has been successfully tested in plants, mainly in monocotyledon species. In this work, we studied the efficiency of SpCas9-NG in the model moss Physcomitrella patens and two Solanaceae crops (Solanum lycopersicum and Solanum tuberosum) for both classical CRISPR-generated gene knock-out and cytosine base editing. We showed that the SpCas9-NG greatly expands the scope of genome editing by allowing the targeting of non-canonical NGT and NGA PAMs. The CRISPR toolbox developed in our study opens up new gene function analysis and plant breeding perspectives for model and crop plants. MDPI 2020-02-04 /pmc/articles/PMC7036883/ /pubmed/32033083 http://dx.doi.org/10.3390/ijms21031024 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Communication
Veillet, Florian
Perrot, Laura
Guyon-Debast, Anouchka
Kermarrec, Marie-Paule
Chauvin, Laura
Chauvin, Jean-Eric
Gallois, Jean-Luc
Mazier, Marianne
Nogué, Fabien
Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title_full Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title_fullStr Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title_full_unstemmed Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title_short Expanding the CRISPR Toolbox in P. patens Using SpCas9-NG Variant and Application for Gene and Base Editing in Solanaceae Crops
title_sort expanding the crispr toolbox in p. patens using spcas9-ng variant and application for gene and base editing in solanaceae crops
topic Communication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7036883/
https://www.ncbi.nlm.nih.gov/pubmed/32033083
http://dx.doi.org/10.3390/ijms21031024
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