Dendrocalamopsis beecheyana var. pubescens is native to the southern provinces of China. In this study, the genetic diversity of 17 accessions from 6 populations of Fujian and Guangxi provinces were assessed. Genetic diversity was estimated using ISSR (inter-simple sequence repeat) markers. Sixteen selected primers produced 185 ISSR loci, of which 167 (90.27%) were found to be polymorphic. Dendrogram analyses revealed that the 17 accessions formed six major groups, whil e the six populations were clustered into three major groups. A high genetic diversity was identified at the species level, with the percentage of polymorphic band (PPB), Nei's gene diversity (He) and Shannon's information index (I) was found to be 90.27%, 0.2497, and 0.3949, respectively. In addition, Nei's differentiation coefficient (Gst) was relatively high (0.5399) and a low rate of gene flow (Nm) of 0.4261 was observed. The results clearly show that there is a high degree of genetic variation at the species level and a low degree of genetic variation at the population level. Moreover, ISSR molecular markers could be effectively utilized for the characterization of D. beecheyana var. pubescens accessions and identification of the relationships within populations.