Plant regeneration from mesophyll protoplasts of tomato cv. Ponderosa

Abstract

Axenic seedlings of tomato cv. Ponderosa were used as material for protoplast culture. Mesophyll protoplasts were treated in an enzyme solution containing 0.5% Cellulase "Onozuka" RS and 0.1% Pectolyase Y-23 for 3.5 hr. at 27°C at 40 spm. Protoplasts were incubated in a liquid medium containing one-fourth strength mineral salts of MS medium(8), organic components of 8 p medium(5), 1.0 mg/l NAA and 0.5 mg/l BA at 29°C in the dark. After the first cell division, the culture was transferred to a light intensity of 300 lux at 25°C. By adding mannitol-free medium stepwise after the first or second cell division, protoplasts were grown to small calluses 0.5-1.0 mm in diameter) by the 30 th day. The calluses, transplanted onto agar medium containing MS basal medium, 1.0 mg/l Zeatin and 0.02 mg/l GA, differentiated adventitious buds within 18 days. Regenerated shoots formed roots on MS agar medium with 0.2 mg/l GA, and developed into fertile plants.