Meta-topolin Improves In Vitro Morphogenesis, Rhizogenesis and Biochemical Analysis in Pterocarpus marsupium Roxb.: A Potential Drug-Yielding Tree

Meta-topolin (mT), a benzyladenine analog [N 6-(3-hydroxybenzylamino) purine] is a highly active cytokinin. The present study evaluates the efficiency of two aromatic cytokinins, mT and BA for inducing in vitro regeneration in a woody legume Pterocarpus marsupium (Roxb.) using cotyledonary node (CN) explants. Of the two cytokinins tested, mT-derived cultures resulted in better shoot multiplication and rhizogenesis than BA. Among the different doses of mT, maximum shoot (9.58 +/- 0.30) induction per explant and average shoot length (4.12 +/- 0.05cm) were recorded on Murashige and Skoog (Physiol Plant 15:473-497, 1962) medium containing 7.5 mu M mT, after 6 weeks of culture. The combined effect of cytokinin and auxin was tested, auxin was mixed with optimum doses of BA or mT separately and the effect of combination was studied. Among the cytokinin-auxin combinations, the highest number of shoots (17.44 +/- 0.25) per explant and average shoot length (5.72 +/- 0.18cm) were achieved on MS medium containing 7.5 mu M mT with 1.0 mu M alpha naphthalene acetic acid in 85% of the cultures after 12 weeks. Meta-topolin alone or in combination with auxin has shown an increase in the quality and number of shoots in comparison to BA. In vitro rhizogenesis in individually regenerated microshoots was carried out on half-strength MS medium augmented with 1.0 mu M indole-3-butyric acid via a two-step procedure method. After 4 weeks, 7.35 +/- 0.11 roots per shootlet with an average root length of 4.54 +/- 0.10cm were recorded in mT-derived microshoots. The well-developed plantlets were acclimatized in a separate batch of single CN explant-derived plantlets. About 80% survival rate was recorded for mT-derived plantlets. Biomass and photosynthetic pigments were also improved in mT-derived plantlets, when compared with the BA derived. Analysis of genetic homogeneity of ten micropropagated plantlets was done through RAPD. Out of 40 RAPD primers, 29 primers produced clearly scorable monomorphic bands, thus exhibiting complete genetic uniformity among in vitro regenerated plantlets.