Facilitating gene editing in potato: a Single-Nucleotide Polymorphism (SNP) map of the Solanum tuberosum L. cv. Desiree genome

François Sevestre,Fabrice Wattebled,Maud Facon,Nicolas Szydlowski

doi:10.1038/s41598-020-58985-6

Abstract

Genome editing is a powerful tool for plant functional genomics allowing for multiallelic targeted mutagenesis. The recent development of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/CRISPR associated protein 9 (Cas9) systems for gene editing in plants allows for simple, cost-effective introduction of site-specific double-stranded DNA breaks. The nuclear genomes of a homozygous doubled-monoploid potato clone (DM) and a heterozygous diploid clone (RH) have been sequenced in 2011. However, common potato cultivars display a highly heterozygous autotetraploid genome thus complicating target design for tetra-allelic gene editing. Here, we report on the SNP physical map of the widely used Solanum tuberosum L. cv. Desiree and on the position of the diverse indels providing an essential tool for target design in genome editing approaches. We used this tool for designing a specific gRNA and successfully knocking-out a newly discovered starch synthase gene (SS6) in potato. Resequencing data are publicly available at the Sequence Read Archive of the NCBI (accession number: PRJNA507597) and will represent a valuable resource for functional genomic studies of various metabolic pathways, cell and plant physiology as well as high-throughput reverse genetics in potato.

Highlights

Genome editing is a powerful tool for plant functional genomics allowing for multiallelic targeted mutagenesis
Zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs) are chimeric proteins composed of a sequence-specific domain fused to a non-specific nuclease (Fok1) that have proven successful in editing the genome of different plant species comprising potato[6,7,8,9]
These data show that polymorphisms occur every 59 bp on average with SNPs being the predominant type of variation (Table 1)

Summary

Introduction

Genome editing is a powerful tool for plant functional genomics allowing for multiallelic targeted mutagenesis. Desiree and on the position of the diverse indels providing an essential tool for target design in genome editing approaches We used this tool for designing a specific gRNA and successfully knocking-out a newly discovered starch synthase gene (SS6) in potato. SNP-based analysis of 23 North American potato modern cultivars showed a population diversity (π) of 0.0105, which was significantly higher than previous resequencing studies of crops[16] Determining both allelic variation of the target region (polymorphisms intrinsic to the studied potato line) and sequence divergence from the DM reference genome is essential for designing targets leading to homozygous gene editing in common tetraploid potato lines. This work represents an essential tool for high throughput functional genomic studies of the potato This was illustrated by the knock-out mutagenesis of SS6, a recently discovered starch synthase isoform most likely involved in starch biosynthesis[2]

Methods

Results

Conclusion