RAD Sequencing of Diverse Accessions of Lepidium campestre, a Target Species for Domestication as a Novel Oil Crop

Lepidium campestre is a wild and winter hardy biennial potential oilseed crop. It can be cultivated as a catch crop and sown in between regular crops. In the Mistra Biotech project, we aim to domesticate L. campestre through a combination of traditional breeding, molecular selection and GM approaches. Target traits are oil quality, oil yield and prevention of premature pod shattering. To provide a molecular toolbox for L. campestre we used Restriction Site Associated DNA (RAD) sequencing to provide a large number of potential single nucleotide polymorphisms (SNP). To get the most informative set of SNPs for both linkage mapping and association studies we selected 8 samples (parents and F1 offspring) from a divergent cross and 10 different accessions for RAD sequencing. DNA was isolated using the protocol by Lopes-Pinto and Vanhala at molmeth.org (2015, http://molmeth.org/protocol/detailed-protocol-isolating-cleaning-and-measuring-dna-lepidium-leaves-rad-sequencing-0).    Single-digest RAD library preparation was carried out using EcoR1 and libraries were sequenced in a single lane of an Illumina HiSeq 2500 using a 125 base paired-end sequencing strategy. RAD reads were analysed de novo using STACKS and SNPs were identified using SAMtools. After filtering, we identified more than 90,000 high quality SNPs that segregate in the experimental cross plus and/or the population samples.  A subset of these SNPs will be selected for a genotyping assay that will enable linkage mapping, QTL mapping, GWAS analyses and also Genomic Prediction in L. campestre.