High-throughput SNP discovery, development and validation of a 30 K target SNP genotyping tool for cultivated flax ( Linum usitatissimu m) breeding and germplasm characterization

The cultivated flax ( Linum usitatissimum L.) is an industrial crop widely cultivated for fiber and seeds, in broad geographical regions around the world. This crop faces many challenges (yield, quality, biotic and abiotic stresses) linked to climate change as almost all other crops. Among solution for improving or breeding new adapted cultivars, marker-assisted selection has been widely applied in plant breeding to enhance crop yield, quality, and tolerance to biotic or abiotic stresses. Recent advance of targeted genotyping-by-sequencing (GBS) offers an ultimate MAS tool to accelerate plant breeding and crop improvement.

To facilitate the utility of SNP-based genotyping, we developed and validated in this study a target SNPs genotyping tool named AT-SNP-30K using Allegro targeted SNPs technology. A total of 41k SNPs were selected from 4.78 million and 3.73 million SNPs identified in two different accessions panels, respectively. Probe design for all these markers was successfully achieved for 35,791 of these markers, representing a 86% conversion success rate. The set of markers was then validated by genotyping a diversity panel comprising 384 individuals, including 376 accessions and eight replicates of the fiber flax Idéo cultivar. The validated genotyping tool includes 35,791 SNPs, covering the fifteen chromosomes with 24,951 high-quality SNPs (MAF> 5%, average low rate of missing data) and 27,247 SNPs having a MAF greater than 1%; demonstrating high polymorphism and excellent genotyping accuracy. The repeatability of genotyping in the validation experiment, reached 99.00% of SNPs for the eight Idéo replicate controls. The AT-SNP-30K genotyping tool is a robust resource for genetic studies, germplasm characterisation and cultivated flax marker-assisted selection studies. It can be used to enhance the breeding of new flax cultivars adapted to the context of climate change.