Callus and azadirachtin related limonoids production through in vitro culture of neem (Azadirachta indica A. Juss)

A protocol was established for the induction of callus and suspension cultures for azadirachtin production from neem explants. Different concentrations and combinations of plant growth regulators (2,4-D, NAA, IAA and BAP) were supplemented in MS medium. Immature flowers, nodular stem sections, leaves immature embryos and mature seeds were used as explants. The highest callus development (78%) was observed when immature flowers were inoculated on MS basal medium with addition of 1.0 mg/l 2,4-D, 1.0 mg/l BAP, 0.2 mg/l NAA and 3% sucrose. The azadirachtin containing liminoids were determined from calli obtained from different explants. Effect of sucrose, glucose, NH(4)NO(3), KNO(3) and urea on cell suspension cultures and azadirachtin contents were also investigated. The dry cell weight and azadirachtin contents increased to 373.1 and 359.2 mu w/ 50 ml when 0.25 and 0.5 g/l NH(4)NO(3) was added in MS liquid media and supplemented with 1.0 mg/l 2,4-D, 0.2 mg/l BAP and 3% sucrose.