ABSTRACT In vitro rhizome production, encapsulation and cold storage of Acorus calamus were attempted for its propagation and ‘true-to-type’ conservation. Shoot cultures were initiated using underground rhizome buds, on 6-benzyladenine (BA) containing Murashige and Skoog (MS) medium. Maximum microrhizome production was observed in presence of 33.3 µM BA, on modified MS medium containing 6% sucrose, 100 mg/L citric acid and 1 g/L polyvinyl pyrrolidone-40. Synthetic seeds were produced from regenerated microtubers by encapsulation in calcium alginate beads. These synthetic seeds were stored in complete darkness at 100C temperature for different durations for mid-term conservation. After cold storage, synthetic seeds were re-cultured in vitro, 100% survival was recorded after the storage of 1, 3 or 6 months; and 80% survival was observed after the storage of 12 months. The microrhizomes were produced roots in 4.9 µM indole-3-butyric acid containing half strength MS medium. All the regenerated plantlets were successfully transferred to field after acclimatization. It is the first report on successful one year in vitro cold storage of A. calamus.