The Japanese serow, Capricornis crispus, is an indigenous bovid species exclusively inhabiting mountain regions in the main Japanese islands, excepting Hokkaido. It had decreased in abundance to its lowest level due to overhunting and deforestation, with its distribution severely fragmented from the middle of the 20th century, many populations of C. crispus currently facing the risk of extinction. The Kii Mountain Range (KM) on Honshu is one such location that has seen a drastic population decline of C. crispus. In this study, we examined genetic characteristics of C. crispus in KM and neighboring regions of the Chubu district, using mtDNA and microsatellite markers, in order to devise strategies for its conservation. Results for mtDNA were characterized by low nucleotide diversity with five endemic and two dominant haplotypes shared by individuals in neighboring regions. A Bayesian skyline plot indicated a gradual increase after the last glacial maximum. For microsatellites, the genetic diversity of C. crispus in KM was comparable to Shizuoka and higher than Shikoku. Recent genetic bottlenecks were strongly suggested in C. crispus in KM. Bayesian clustering showed a genetic cline between KM and neighboring regions, where multivariate analysis suggested three local populations. A Mantel test indicated male-biased dispersal. These results indicate that C. crispus in KM and neighboring regions constitute multiple local populations, connected through restricted gene flow. For the conservation of C. crispus, it is important to define small-scale conservation units, among which genetic connectivity should be facilitated to prevent further loss of genetic diversity.