An Efficient Protocol for Cyclic Somatic Embryogenesis in Neem (Azadirachta indica A Juss.)

Neem is a highly heterozygous and commercially important perennial plant. Conventionally, it is propagated by seeds which loose viability within two weeks. Strictly cross pollinating nature of the plant causes serious barrier to the genetic improvement by conventional methods. Alternative methods of tree improvement such as somatic hybridization, mutagenesis and genetic transformation require an efficient in vitro plant regeneration system. In this regard, somatic embryogenesis particularly secondary somatic embryogenesis may offer an effective system for large scale plant propagation without affecting the clonal fidelity of the regenerants. It can be used for synthetic seed production, which further bolsters conservation of this tree species which is otherwise very difficult The present report describes the culture conditions necessary to induce and maintain repetitive somatic embryogenesis, for the first time, in neem. Out of various treatments tested, the somatic embryos were induced directly from immature zygotic embryos of neem on MS + TDZ (0.1 μM) + ABA (4 μM), in more than 76 % cultures. Direct secondary somatic embryogenesis occurred from primary somatic embryos on MS + IAA (5 μM) + GA3 (5 μM) in 12.5 % cultures. Embryogenic competence of the explant as well as of the primary embryos was maintained for a long period by repeated subcultures at frequent intervals. A maximum of 10 % of these somatic embryos were converted into plantlets. Keywords—Azadirachta indica A. Juss., Cytokinin, Somatic embryogenesis, zygotic embryo culture. Abbreviations—ABA abscisic acid; BAP 6benzylaminopurine; CH casein hydrolysate; 2,4-D 2,4dichlorophenoxyacetic acid; GA3 gibberellic acid; IAA indoleacetic acid, IBA indole-3-butyric acid; Kn N6furfuryladenine, NAA -naphthalene acetic acid.