Identifying Genes Conferring Rust Resistance in Adzuki Bean (Vigna angularis) Through Genome-Wide Association Analysis Based on Genotyping-by-Sequencing (GBS)

Adzuki bean (Vigna angularis) is an important pulse crop, and rust disease severely restricts its production. Identifying resistance genes is essential for accelerating the development of rust-resistant cultivars through marker-assisted breeding. In this study, genotyping-by-sequencing (GBS) was performed on 178 adzuki bean accessions, generating 124.87 Gb of sequencing data and yielding 12,153 high-quality single-nucleotide polymorphism (SNP) markers with an average density of 26.56 SNPs/Mb across the genome. Principal component analysis (PCA) and K-means clustering divided the 178 accessions into three distinct genetic clusters, with the first five principal components explaining 31.90% of the total genetic variation. Population structure analysis based on these SNPs revealed that the cross-entropy criterion reached its minimum at K = 7, partitioning the panel into seven genetic subpopulations (C1–C7). A neighbor-joining phylogenetic tree further resolved the accessions into five major groups. Rust resistance was assessed at the seedling stage under greenhouse conditions with artificial inoculation of Uromyces vignae isolate ZXL01, and at the adult stage under natural field infection at a single experimental site. Genome-wide association analysis (GWAS) was conducted separately for the two assessments and identified fifteen SNPs significantly associated with rust resistance. Ten SNPs were identified at the seedling stage on chromosomes 1 and 3, while the remaining five SNPs were detected at the adult stage on chromosome 9. The SNPs associated with adult-stage rust resistance were converted into Kompetitive Allele-Specific PCR (KASP) markers, which were subsequently validated in an independent panel. The validation confirmed that the resistance-associated alleles were predominantly detected in highly resistant accessions, suggesting that these markers have potential utility for the preliminary screening of rust-resistant germplasm, although further breeding validation is required.