Genome-Wide Discovery of SSR Markers Based on Whole-Genome Resequencing Data of Dendrobium officinale

<i>Dendrobium officinale</i>, a perennial epiphytic herb of the orchid family renowned for its ornamental value and diverse medicinal properties, has attracted considerable global attention due to its rarity and increasing market demand. However, large-scale cultivation has led to confusion regarding its germplasm resources and genetic backgrounds, posing significant challenges for the effective conservation, management, and utilization of <i>D. officinale</i> germplasm. In this study, we systematically analyzed the abundance and characteristics of simple sequence repeats (SSRs) and developed highly polymorphic genomic SSR markers using whole-genome resequencing (WGRS) data from 15 <i>D. officinale</i> genotypes. A total of 494,783 SSRs were identified in the "Niu2020" reference genome. Mononucleotide repeats were the most abundant motifs, followed by dinucleotide repeats, with A/T and AT/TA being the predominant types, respectively. Of the SSRs with unique flanking sequences (unique SSRs), 121,544 out of 388,553 (31.28%) were polymorphic across the 15 genotypes. The polymorphism information content (PIC) values of these SSRs ranged from 0.13 to 0.92, with an average of 0.59. Subsequently, 10,364 pairs of SSR primers were successfully designed from polymorphic SSR loci with PIC values ≥ 0.5 and major allele size differences ≥ 3 bp. Ultimately, 20 primer pairs were randomly selected for experimental validation. Of these, 18 successfully amplified the target fragments and exhibited high levels of polymorphism (PIC values ranged from 0.31 to 0.74), confirming the effectiveness and practical utility of the newly developed markers. The SSR fingerprints and polymorphic markers developed in this study provide a valuable resource and establish a robust technical foundation for enhancing the efficiency of cultivar identification, genetic research, and molecular breeding in <i>D. officinale</i>.