In vitro propagation, in vitro flowering, ex vitro root regeneration and foliar micro-morphological analysis of Hedyotis biflora (Linn.) Lam

Abstract

Hedyotis biflora (Linn.) Lam is an important herb, used in the Indian and Chinese traditional systems of medicines. Indiscriminate harvesting of this plant resulted in decline of its population from the natural habitats. The present study reports effective micropropagation and in vitro flowering in H. biflora. Healthy plants were maintained in the greenhouse to collect fresh explants (nodal segments). Average 3.2 shoot bud/node with 88% shoot regeneration obtained on Murashige and Skoog’s (MS) medium with 2.0 mg L−1 6-benzylaminopurine (BAP) and additives like 50 mg L−1 of ascorbic acid and 25 mg L−1 each of arginine, adenine sulphate and citric acid. About 84.5 shoots, with 8.1 cm shoot length were obtained on MS basal medium supplemented with 1.0 mg L−1 BAP, 0.5 mg L−1 kinetin (Kin) and 0.1 mg L−1 indole-3 acetic acid (IAA) and additives. In vitro flowers were induced in 91% cultures (6.8 flowers per shoot) when the shoots were cultured on MS medium supplemented with 1.5 mg L−1 BAP and 0.1 mg L−1 IAA at 12-h photoperiods. In vitro raised shoots were rooted with the help of indole-3 butyric acid (IBA) via in vitro and ex vitro techniques. The micropropagated plants were acclimatized with 91% survival rate in the field. Upon transfer of plantlets from the in vitro to ex vitro environments, there were significant changes in stomata, vein-islets and raphids. These micro-morphological changes may help micropropagated plantlets to establish under the field conditions. The present protocol is suitable for large scale micropropagation, conservation and commercial production of this plant. The protocol of in vitro flowering could be useful in understanding the in vitro pollination biology of H. biflora.