Genetic diversity and population structure analysis in wood-apple (Limonia acidissima L.) using simple sequence repeat (SSR) markers

Wood-apple (Limonia acidissima L., Rutaceae) is an underutilized fruit tree species in the Indian subcontinent. The edible fruit and other parts of the wood-apple tree have immense medicinal and nutritional properties. In this study, we used 24 polymorphic simple sequence repeats (SSRs), developed de novo through RNA sequencing, to estimate the genetic diversity and population structure in 96 wood-apple samples collected from 16 states of India. We identified 11,401 SSRs from 215,859 unigenes. The dinucleotide repeat motifs were the most abundant SSRs (56.89%), followed by trinucleotide (37.61%) and tetranucleotide repeats (3.63%). SSR profiling detected 129 alleles, with an average of 5.37 per locus and an average PIC value of 0.62. The Jaccard's genetic dissimilarity among 96 samples ranged from 0.39 to 1.0 (average 0.94 & PLUSMN; 0.03). Cluster analyses by Neighbour-joining (NJ) and principal coordinates analysis (PCoA) grouped all samples into three clusters. The Bayesian model-based structure analysis also identified three subpopulations. Population genetic analysis showed moderate gene diversity (0.226 & PLUSMN; 0.007) and Shannon's information index (0.369 & PLUSMN; 0.009) in the species. Analysis of molecular variance (AMOVA) revealed higher genetic variance within the populations than among populations, while the F-ST statistics (0.134, p < 0.010) indicated low but significant genetic differentiation. The overall results indicated moderate to high genetic diversity among wood-apple samples. This study identified 14 diverse samples and nine highly informative SSRs that could be useful for developing breeding programs to improve the species and conserve its genetic resources.