Shorea obtusa is a keystone species of the dry deciduous dipterocarp forest in Thailand. In this study, the genetic structure and diversity of this species were evaluated by means of five microsatellite markers. A total of 146 trees were collected from five populations encompassing major forest regions of Thailand. High levels of genetic diversity were found among the five populations with the average He of 0.664. Genetic differentiations between populations, although significant, were low with approximately 3% of genetic variation partitioned among populations. This may indicate that the populations sampled were recently part of a continuous population. A tree constructed using the unweighted pair group method with arithmetic average, based on Nei‘s genetic distance, divided the populations into three groups. This separation was consistent with the altitudinal zonation of the populations, thus indicating that altitude might play a significant role in the genetic structure of S. obtusa. Areas of high genetic diversity were identified which could be considered priorities for conservation.