Genotyping Brassica Crops using SNP Array

Genotyping for association mapping was performed using the Brassica 60 K Illumina® Infinium SNP array [32 (link)] according to the manufacturer’s protocol (https://www.illumina.com/techniques/microarrays.html) in the National Key Laboratory of Crop Genetic Improvement, National Subcenter of Rapeseed Improvement in Wuhan, Huazhong Agricultural University, 430070 Wuhan, China. Illumina BeadStudio genotyping software was used for SNP data clustering and calling according to a previously described protocol [31 (link)]. SNPs with a call frequency of <0.9 and a minor allele frequency (MAF) of ≤0.05 were excluded in this research. The remaining SNPs were scrutinized visually and those SNPs that were resolved as three clearly separated clusters (AA, AB, and BB) in the tested B. napus material were used for further research. In addition, to identify the physical position of SNP markers, the source sequences for designing SNP probes of the Brassica 60 K SNP arrays were used to perform a BlastN search against the B. napus ‘Darmor-bzh’ reference genome (version 4.1, http://www.genoscope.cns.fr/brassicanapus/data/) [26 (link)]. Only the top BLAST hits (E values of < “1.0 E⁻¹⁰”) were considered to be mapped in the genome, while BLAST matches to multiple loci with the same top identity were not considered to be mapped [31 (link), 35 ].