Microcallus formation from Hevea brasiliensis protoplasts isolated from embryogenic callus.

Conditions for successful culture of rubber tree (Hevea brasiliensis) protoplasts were investigated. Protoplasts, derived from embryogenic callus, regenerated cell walls then underwent division when embedded in alginate and cultivated on a modified Murashige and Sook medium (9 μM 2,4-dichlorophenoxyacetic acid, 0.6 M glucose, 0.93 μM kinetin, NH 4 (+) reduced by half) in the presence of nurse cells (tobacco feeder cell layer). The presence of nurse cells was essential to maintain viability and sustain protoplast division. Several parameters which influenced the plating efficiency were analysed, such as the density of feeder cells and the duration of contact of the feeder layer.