Plant regeneration through somatic embryogenesis and RAPD analysis of regenerated plants in Tylophora indica (Burm. f. Merrill.)

A procedure for the regeneration of complete plantlets of Tylophora indica, from cultured leaf Callus via somatic embryogenesis is described. Callus i nduction from leaf explants was on Murashige and Skoog (MS) medium with dif ferent concentrations of 2,4-dichlorophenoxy-acetic acid (2,4-D; 0.0-3 mg l (-1); 0.0-13.56 muM) and kinetin (Kn; 0-01 mg l(-1); 0.05 muM). The best re sponse for callus induction was obtained on MS medium containing 2 mg l(-1) (9.04 muM) 2,4-D and 0.01 mg l(-1) (0.05. muM), Kn. After two subcultures on the same medium the embryogenic Callus was transferred to MS medium with different concentrations of the cytokinin, 6-benzyladenine (0.5-3 mg l(-1) ; 2.22-13.32 muM) and 2-isopentenyladenine (2ip; 0.53 mg l(-1); 2.46-14.76 muM) along with 0.01 mg l(-1) (0.05 muM) indole-3-butyric acid (IBA) for so matic embryo development and maturation. MS medium with 2 mg l(-1) (9.84 mu M) 2ip produced the maximum number of mature somatic embryos. The mature em bryos were bipolar and on transfer to MS basal medium produced complete pla ntlets. After hardening the regenerants were planted in the Gudalur forests of Western Chats. Total DNA was extracted from 14 regenerants and the moth er plant. Random amplified polymorphic DNA (RAPD) analysis was carried out using 20, arbitrary oligonucleotides. The amplification products were monom orphic among all the plants revealing the genetic homogeneity and true-to-t ype nature of the regenerants.