Improved procedure for protoplast culture and plant regeneration of spinach (Spinacia oleracea L)

Protoplasts of Spinacia oleracea L. were cultured in a modified half-strength MS medium supplemented with KM8p vitamins, 1.0 mg . liter(-1) BA, 1.0 mg . liter(-1) 2,4-D, and glucose or mannitol. Plating efficiency in the medium containing glucose as an osmoticum was much higher than that in the medium containing mannitol, The addition of KM8p organic acids to the glucose medium promoted the division of protoplasts slightly; the same addition to a medium containing mannitol appeared inhibitory to the cell division. Several shoots were formed from calli obtained on the MS medium containing 1.0-10.0 mg . liter(-1) IAA or IBA. Adventitious root was developed most vigorously on the MS medium containing 2% sucrose, 0.5 mg . liter(-1) IBA, and 0.8% agar when pH was adjusted to 6.3. Some regenerated plantlets transferred to a greenhouse produced many seeds.