A blueprint for gene function analysis through Base Editing in the model plant Physcomitrium (Physcomitrella) patens
Guyon-Debast, Anouchka (Institut Jean-Pierre Bourgin)
Alboresi, Alessandro (University of Padova)
Terret, Zoé (Institut national de recherche pour l'agriculture, l'alimentation et l'environnement)
Charlot, Florence (Institut Jean-Pierre Bourgin)
Berthier, Floriane (Institut Jean-Pierre Bourgin)
Vendrell-Mir, Pol (Centre de Recerca en Agrigenòmica)
Casacuberta i Suñer, Josep M. 1962- (Centre de Recerca en Agrigenòmica)
Veillet, Florian (Institut de Génétique, Environnement et Protection des Plantes)
Morosinotto, Tomas (University of Padova)
Gallois, Jean-Luc (Institut national de recherche pour l'agriculture, l'alimentation et l'environnement)
Nogué, Fabien (Institut national de recherche pour l'agriculture, l'alimentation et l'environnement)

Fecha:	2021
Resumen:	CRISPR-Cas9 has proven to be highly valuable for genome editing in plants, including the model plant Physcomitrium patens. However, the fact that most of the editing events produced using the native Cas9 nuclease correspond to small insertions and deletions is a limitation. CRISPR-Cas9 base editors enable targeted mutation of single nucleotides in eukaryotic genomes and therefore overcome this limitation. Here, we report two programmable base-editing systems to induce precise cytosine or adenine conversions in P. patens. Using cytosine or adenine base editors, site-specific single-base mutations can be achieved with an efficiency up to 55%, without off-target mutations. Using the APT gene as a reporter of editing, we could show that both base editors can be used in simplex or multiplex, allowing for the production of protein variants with multiple amino-acid changes. Finally, we set up a co-editing selection system, named selecting modification of APRT to report gene targeting (SMART), allowing up to 90% efficiency site-specific base editing in P. patens. These two base editors will facilitate gene functional analysis in P. patens, allowing for site-specific editing of a given base through single sgRNA base editing or for in planta evolution of a given gene through the production of randomly mutagenised variants using multiple sgRNA base editing.
Derechos:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, i la comunicació pública de l'obra, sempre que no sigui amb finalitats comercials, i sempre que es reconegui l'autoria de l'obra original. No es permet la creació d'obres derivades.
Lengua:	Anglès
Documento:	Article ; recerca ; Versió publicada
Materia:	Adenine deaminase ; APRT ; Base editing ; Cas9 ; CRISPR ; Cytosine deaminase ; Physcomitrella patens ; Physcomitrium patens
Publicado en:	The new phytologist, Vol. 230, Issue 3 (May 2021) , p. 1258-1272, ISSN 1469-8137

DOI: 10.1111/nph.17171
PMID: 33421132

15 p, 2.4 MB

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