Title :

The Solanum tuberosum GBSSI gene: a target for assessing gene and base editing in tetraploid potato

Author(s) :

Veillet, Florian [Auteur]
Chauvin, Laura [Auteur]
Kermarrec, Marie-Paule [Auteur]
Sevestre, Francois [Auteur]
Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF]
Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - UAR 3290 [MSAP]
Merrer, Mathilde [Auteur]
Terret, Zoé [Auteur]
Szydlowski, Nicolas [Auteur]
Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF]
Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - UAR 3290 [MSAP]
Devaux, Pierre [Auteur]
Gallois, Jean-Luc [Auteur]
Chauvin, Jean-Eric [Auteur]

Journal title :

Plant Cell Reports

Abbreviated title :

Plant Cell Rep

Volume number :

38

Pages :

1065-1080

Publisher :

Springer Nature

Publication date :

2019-05-17

English keyword(s) :

Genome editing
CRISPR-Cas9
Cytidine base editor
Potato
GBSS
HRM

HAL domain(s) :

Sciences du Vivant [q-bio]
Chimie/Chimie théorique et/ou physique

English abstract : [en]

Genome editing has recently become a method of choice for basic research and functional genomics, and holds great potential for molecular plant-breeding applications. The powerful CRISPR-Cas9 system that typically produces ...
Show more >Genome editing has recently become a method of choice for basic research and functional genomics, and holds great potential for molecular plant-breeding applications. The powerful CRISPR-Cas9 system that typically produces double-strand DNA breaks is mainly used to generate knockout mutants. Recently, the development of base editors has broadened the scope of genome editing, allowing precise and efficient nucleotide substitutions. In this study, we produced mutants in two cultivated elite cultivars of the tetraploid potato (Solanum tuberosum) using stable or transient expression of the CRISPR-Cas9 components to knock out the amylose-producing StGBSSI gene. We set up a rapid, highly sensitive and cost-effective screening strategy based on high-resolution melting analysis followed by direct Sanger sequencing and trace chromatogram analysis. Most mutations consisted of small indels, but unwanted insertions of plasmid DNA were also observed. We successfully created tetra-allelic mutants with impaired amylose biosynthesis, confirming the loss of function of the StGBSSI protein. The second main objective of this work was to demonstrate the proof of concept of CRISPR-Cas9 base editing in the tetraploid potato by targeting two loci encoding catalytic motifs of the StGBSSI enzyme. Using a cytidine base editor (CBE), we efficiently and precisely induced DNA substitutions in the KTGGL-encoding locus, leading to discrete variation in the amino acid sequence and generating a loss-of-function allele. The successful application of base editing in the tetraploid potato opens up new avenues for genome engineering in this species.Show less >

Language :

Anglais

Peer reviewed article :

Oui

Audience :

Internationale

Popular science :

Non

Administrative institution(s) :

Université de Lille
CNRS

Collections :

• Unité de Glycobiologie Structurale et Fonctionnelle (UGSF) - UMR 8576

Research team(s) :

Plant Storage Polysaccharides

Submission date :

2020-12-14T14:41:35Z
2021-01-05T14:07:21Z