Cell regeneration and sustained division of protoplasts from cotton (Gossypium hirsutum L.)

Protoplasts were isolated from 12 day old subcultured phytohormone habituated callus tissue of Gossypium hirsutum L. (0.5% cellulysin-Calbiochem, 0.6% macerase-Calbiochem, 0.7M mannitol, and pH 5.0). After separation and purification (0.35M sucrose floatation medium), the protoplasts were cultured (K3 media of Kao et al., 1974 with 0.9 μM BAP, 5 μM IAA and 0.35M sucrose) in both liquid and solid medium at a density of 5×10(5) protoplasts/ml. Four weeks after isolation, cell regeneration and callus formation was observed.