Transcriptome-derived SSR markers uncover ornamental trait loci and establish a core collection for Rhododendron breeding

Rhododendron spp. ranks among China’s top ten traditional ornamentals, yet cultivar innovation remains limited by germplasm redundancy. We generated a multi-tissue transcriptome of Rhododendron × pulchrum, identified 5273 unigenes that contained SSR motifs, and developed 30 polymorphic EST-SSR markers (PIC = 0.56). These loci amplified 3–18 alleles each and displayed transferable amplification across three subgenera. Genotyping of 141 accessions revealed high diversity (Ho = 0.40, He = 0.60, I = 1.31) and six well-supported genetic clusters (STRUCTURE, NJ). An exploratory association mapping based on GLM and MLM detected eight SSR loci (A16, A272, A313, A345, A349, A353, A362, A363) significantly linked to flowering time, flower form and plant habit (P 0.05), confirming its representativeness. These resources-the first EST-SSR kit with cross-subgenus transferability, putative trait-linked markers, and a validated core collection-provide a theoretical framework and foundational molecular resources for future genetic dissection and molecular breeding in Rhododendron.