[Objective] This study aimed to investigate the genetic variation of wild and cultivated populations of Castanopsis hystrix. [Method] Genetic variation of five wild populations and three cultivated populations of Castanopsis hystrix, was investigated with ISSR-PCR amplification. Totally, 151 individuals were selected and analyzed by amplification using nine pairs of ISSR primers screened. [Result] Each primer pair produced 7-20 bands and 122 polymorphic bands were obtained. At population level, ISSR diversity in the wild populations (P=59.84%, HPOP=0.182 7, I=0.285 6) was higher than which in cultivated ones (P=54.87%, HPOP=0.136 6, and I=0.219 8). The genetic differentiation coefficient among wild populations (GST) was 0.99. The similar population structure was found in three cultivated populations (GST=0.127 5). According to the UPGMA cluster analysis, the genetic distance among wild populations became larger with the increase of geographical distance. [Conclusion] Compared with other seed plants, with either a similar life history or various breeding system attributes, relatively low level of genetic diversity was observed in these five wild populations, which was caused by population size reduction and habitat fragmentation related to human activities. The formation of population structure may be explained by the species’ breeding system.