The present investigation was undertaken to study establishment of clean cultures, shoot multiplication and short and long term in vitro conservation of shoot tips of Rauvolfia serpentina L. The present study was aimed to standardize an efficient protocol for in vitro multiplication and conservation of Rauvolfia serpentina L. Simple surface sterilization was found effective to establish clean cultures. In vitro culture establishment study indicated the importance of surface sterilization treatment in maintaining disease free cultures in tissue culture. In vitro multiplication is a pre-requisite for in vitro conservation. A combination of auxin (NAA) and cytokinin (BA) in MS medium were studied. Shoot multiplication was found better in MS medium containing both NAA and BA as compared to BA alone. So the present study revealed the importance of growth regulator during shoot regeneration. In vitro conservation study revealed the feasibility of using encapsulated shoot tips for conservation purpose. The encapsulated shoot tips of R. serpentina L. could be conserved well at culture room condition (25°C ± 2°C, 16 hrs photoperiod). The encapsulated shoot tips exhibited best response of regrowth in MS medium supplemented with NAA and BA as compare to BA alone and hormone free medium. Using cryopreservation technique success was achieved in obtaining regrowth in control (without LN2 treated cultures). Using encapsulation dehydration technique, it was observed that there was negative correlation in moisture content of beads was reduced from 80-100 per cent during 0-4 hours air desiccation, the growth rate of controls was increased. Using vitrification technique, shoot tips treated with high sucrose and PVS2 (cryoprotectant) exhibited better survival and regrowth in controls (without LN2 treated cultures), compare to without sucrose and PVS2 treated cultures. This study showed the importanc of dehydrating agents and cryoprotectants in cryopreservation technique.