Glehnia littoralis Fr. Schmidt et Miquel is an important medicinal plants in East Asian countries. This plant species naturally distributed in Korea, Japan, China, and Taiwan, but it is a rare plants living in the coastal dune in Korea. To investigate the genetic variation, genetic diversity and genetic evolutionary relationships of 14 different geographical G. littoralis, ITS sequence and random amplified polymorphic DNA (RAPD) were analyzed. On the basis of ITS sequences, it was clearly showed that the ITS1 and ITS2 sequences among 14 populations are identical regardless of geographical origin excepting 2 bp in pair-wise comparison of ITS1. Furthermore, RAPD results also showed that 14 different geographical G. littoralis produce various polymorphic patterns without critical relationship among neighboring regions. These combined results suggest that the geographical variation and genetic evolution of G. littoralis is stable and provide important information on genetic diversity, and conservation of this rare plant species in situ and ex situ.