メロン子葉プロトプラストからの不定芽再分化

Abstract

Protoplasts, isolated from cotyledons of aseptically germinated seeds of the melon 'Charen-tais', were used to investigate the effects of hormones and osmotic conditions on cell divi-sion and plantlet regeneration.1. Division of protoplasts occurred at a high frequency during the initial incubation peri-od on the modified MS medium (200 mg•liter-1 NH4NO3 and 1% sucrose) containing a wide range of 2, 4-D concentrations. Protoplast division was continuous in a medium containing 0.05 mg•liter-1 2, 4-D and 0.5 mg•liter-1 BA even as the protoplasts mature and formed cell walls.2. Three osmotic conditions; mannitol only and an equimolor conc. of mannitol and sucrose and that of mannitol and glucose were used to clarify the role of the osmoticum on the shoot regeneration from calli. No obvious difference among the three osmotic conditions was ob-served in the frequency of cell division at initial culture and in callus formation. The callus grown in a medium containing mannitol and glucose differentiated shoots at the highest fre-quency among the media tested.3. More shoots were regenerated in a MS medium containing 0.5 mg•liter-1 GA3 and 2 mg•liter-1 BA than were formed with a combination of GA3 and zeatin. The GA3 com-bined with cytokinins promoted more shoots formation from calli than did the IAA-cytokinin combinations.The following protocol for shoot regeneration of melon was quite efficient: a) Culture the protoplasts in the modified MS medium supplemented with 0.05 mg•liter-1 2, 4-D, 0.5 mg•liter-1 BA, 0.2 M mannitol and 0.2 M glucose. b) Transfer the callus grown in the above medium to regeneration medium supplemented with 2 mg•liter-1 BA and 0.5 mg•liter-1GA3. On the average, three shoots per callus were regenerated.Many regenerated shoots may appear abnormal and will not elongate normally, in which case, transplant the explant intact onto a MS medium containing 0.1 mg•liter-1 BA and 0.2 mg-liter-1 GA3. Subculture the explant several times to promote elongation of the shoots. Finally, subculture the elongated shoot on MS agar medium with 1 mg•liter-1 NAA for 24 hr and then transfer it to one without any hormone. The treated shoot will regener-ate roots and grow into a normal plant.