In vitro microrhizomes induction and genetic stability of a medicinal plant Acorus calamus L. towards germplasm conservation through synthetic seed production

Germplasm conservation through in vitro multiplication is one of the important mass cloning method to protect endangered medicinal plants. Synthetic seed technology offers potential advantages by increased efficiency for in vitro propagation in terms of space, labour, time and overall experimental costs. In vitro shoot formation (3.1±0.34 shoots per in vitro raised explants with shoot length of 2.9±0.11 cm of Acorus calamus (Bach) was achieved after 4 weeks of culture in Murashige & Skoog (MS) media supplemented with 4 mg l-1 BAP and 1 mg l-1 IAA with 30g sucrose l-1. Highest number of shoot multiplication per explant was recorded in 8 mg l-1 BAP and 2mg l-1 IAA and successful profuse rooting was observed in 1 mg l-1 IBA (3.6±0.2 roots per explant) with an average root length of 4.7±0.6 cm. Synthetic seeds of in vitro raised microrhizomes were produced in a artificial matrix containing 3% (w/v) sodium alginate prepared in MS basal medium for 20 min and dropped in to 3% (w/v) calcium chloride solution for 30 min for polymerization. The encapsulated microrhizomes were stored at cold (10oC) in dark condition with different storage intervals of 3 and 6 months to evaluate the effect of storage duration and germination of zygotic embryos in A. calamus. From the RAPD analysis of both in vivo and in vitro regenerated plants it was observed that there were no significant variation of DNA profiles, thus ensuring the genetic stability of the plants and regeneration of true to type plants.