Abstract
An efficient and rapid protocol has been reported for the first time for plant regeneration through somatic embryogenesis from corm and young corm buds of Iphigenia indica Kunth et Benth, an important colchicine yielding threatened medicinal plant of Colchicaceae. Soft, friable and well proliferating callus was initiated in Murashige and Skoog's modified basal medium (MS) supplemented with 3% (w/v) sucrose, 0.05 mg/l (w/v) each of nicotinic acid, thiamine-HCl, pyridoxine-HCl, ascorbic acid, 0.1 mg/l (w/v) glutamine, 0.25% (w/v) Gelrite® and a combination of α-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP) (2.0/0.5 mg/l). Such callus became nodular and embryonic in nature after transferring to the same MS modified medium containing 2,4-dichlorophenoxyacetic acid (2,4-D) (1.0 mg/l). The globular embryos were observed in this medium after 4 weeks in culture. The transition of globular stage occurred when embryonic callus mass was shifted to half strength of MS medium containing 0.1 mg/l kinetin with production of maximum percentage of embryos showing distinct epicotyl and hypocotyl differentiation (65.4±0.46). These growing embryos developed into complete plants with well-formed root and shoot systems in the same medium containing 1% (w/v) sucrose. All the regenerates showed stability in chromosome number (2n=22) with 87.4% survival rate after 2–3 weeks of hardening. High Pressure Liquid Chromatography (HPLC) analysis revealed higher amount of colchicine in the regenerated corm-like bulbous structures than in vivo corms. However, both regenerated and in vivo shoots contained negligible amount of colchicine.
