Utilization of plant tissue culture technique for mass propagation and conservation of endangered medicinal plants

We established protocols for mass propagation of three endangered medicinal plants, Rauvolfia serpentina, Gloriosa superba, and Smilax zeylanica through in vitro shoot tip culture. Apical and axillary buds of young sprouts from selected plants were used as explants. For shoot induction in R. serpentina and S. zeylanica MS basal medium, supplemented with 1.5 mg/l BAP + 0.5 mg/l NAA and 1.5 mg/l BAP + 0.2 mg/l NAA was required, in which 95 and 92% cultures produced shoots with 3.9 and 6.1 shoots per culture, respectively, whereas for Gloriosa superba, 135 basal medium with 0.5 mg/l BAP + 0.5 mg/l Kn + 0.2 mg/l NAA was needed, in which 82% cultures produced shoots with 4.4 shoots per culture. Repeated subcultures in the same nutrient medium mentioned above, resulted in rapid shoot multiplication with 25(1) and 10 shoots per culture of R. serpentina and S. zeylanica, respectively but in G. superba the number of shoots was not found to increase even through several subcultures. For further development of the medium, casein hydrolysate (CH) (50 - 200 mg/l) and coconut milk (CM) (5 - 20%) were added individually or simultaneously to the medium for all the three species but their effects were not found to be satisfactory for the culture R. serpentina. In G. superba culture addition of CH and CM individually or simultaneously was not so effective; only growth of individual shoots increased in length with 100 mg/l CH, whereas in S. zeylanica addition of 15% CM increased the number of shoots up to 15 per culture. In vitro raised shoots rooted on half strength MS medium with auxin. Concentration and combination of auxins were found to be species specific, which were 1.0 mg/l each of IBA and IAA for R. serpentina, 0.5 mg/l each of IBA and NAA for G. superba and 1.0 mg/l NAA + 0.5 mg/l IAA for S. zeylanica. For acclimatization and transplantation, the plants in the rooting culture tubes were kept in normal room temperature for 7 days before transplanting in pots where plants were reared for three weeks. The survival rate of regenerants of G. superba were found to be 42%, whereas those of other two species were 85 - 90%. The technique described here may be a promising method for propagation as well as for sustainable use and conservation.