Efficient in vitro regeneration of Leucaena leucocephala using immature zygotic embryos as explants

A method for efficient in vitro regeneration of Leucaena leucocephala cv. K636 was developed using immature zygotic embryos as the explant. Multiple shoot induction was achieved by culturing the explants on half-strength Murashige and Skoog (MS) medium supplemented with thidiazuron (TDZ) (0.03 to 1.5 mu M). The maximum number of shoots per explants was achieved in 0.26 mu M TDZ-supplemented media. TDZ concentration significantly influenced the induction of multiple shoots as well as the shoot-length. TDZ at 0.35 mu M or higher concentrations resulted in abnormal stunted shoots. Full-strength MS media devoid of any plant growth regulator were used for shoot elongation. In vitro root induction was achieved in half-strength MS media supplemented either with 0.54 mu M 1-naphthaleneacetic acid (NAA) or with 14.76 mu M indole-3-butyric acid (IBA) in combination with 0.23 mu M kinetin (Kn). Media supplemented with 14.76 mu M IBA in combination with 0.23 mu M Kn induced twofold-threefold more roots than media supplemented with 0.54 mu M NAA. However, the average root length was lower in 14.76 mu M IBA in combination with 0.23 mu M Kn than in 0.54 mu M NAA. Regenerated plants were maintained under normal condition after hardening. Plants, which were rooted on media supplemented with 14.76 mu M IBA in combination with 0.23 mu M Kn showed higher survival rate during hardening than those rooted on 0.54 mu M NAA supplemented media.