LAUSR

ABSTRACT Expressed sequence tagged (EST) microsatellite (SSR) markers were newly developed for Callicarpa subpubescens, a species endemic to the Bonin (Ogasawara) Islands of Japan, to investigate gene flow, genetic structure, and evolution of the species. A total of 87,334 contigs derived from de novo sequencing were obtained from which 1,826 primers bordering candidate SSRs were designed. Success of amplification and presence of polymorphisms were tested for each primer pair using two populations of C. subpubescens, and one population of C. parvifolia closely related to C. subpubescens to check cross-species amplification. Twenty-nine of the 48 primer pairs showed clear amplification, with 27 showing polymorphisms in the tested populations. For the 27 polymorphic loci, across the two species, the number of alleles ranged from two to 23 (average = 5.93), and expected heterozygosity per locus were 0.05–0.91 (average = 0.52). Bayesian clustering analysis for C. subpubescens showed that the most likely number of clusters is two, with each population dominated by a unique cluster. These new EST-SSR markers presented here can be used for population genetic study of C. subpubescens and its congener C. parvifolia.