【摘要】 从豇豆（Vigna Sinensis Endl.）未成熟子叶分离原生质体,纯化后的产量为1.5×10⁶-2×10⁶/g fr.wt。在MS、B₅和KM_8P 3种培养基中,豇豆未成熟子叶原生质体培养以MS为优,细胞分裂频率为25.1%-30.7%。培养20—30天后,形成肉眼可见的小愈伤组织。在MS液体培养基中形成的愈伤组织转移到MSB（即MS无机盐+B₅维生素）+2 mg/L2,4-D+0.5mg/L BA培养基上继代培养,获得胚性愈伤组织。将胚性愈伤组织转移到MSB+1.0 mg/L 2,4-D+0.25 mg/L BA液体培养基中建立悬浮培养。随后,悬浮培养的胚性愈伤组织再转移到MSB+0.1 mg/L IAA+0.5 mg/L KT琼脂培养基上,经光照培养7-10天后,可观察到大量体细胞胚的形成。部分体细胞胚可发育到子叶期。子叶胚转入新鲜的MSB+0.5 mg/L KT+0.1 mg/L IAA培养基后,迅速萌发生长成具真叶的小植株或小苗。更多还原

【Abstract】 Immature cotyledons of cowpea （Vigna sinensis Endl。） were used for protoplast isolation. Enzyme solution for protoplast isolation contained 4% cellulase Onozuka R-10,0.3% Macerozyme R-10 and 2% hemicellulase. The purified protoplasts were cultured in B₅,MS or KM^8P liquid medium in dark （25℃） at a density of 1 × 10⁵ —5 × 10⁵/ml. The protoplasts started cell division in 3—5 days . Sustained cell divisions resulted in formation of cell clusters and small calli,with cell division frequency reaching 23% —28% in MS medium . Calli of 2 mm in size were transferred onto MSB （MS salts+B₅ vitamins） medium with 2 mg/L 2,4-D, 0. 5mg /L BA forfurther growth. Embryogenic calli appeared on this medium. After passage to fresh medium with the same composition, the embryogenic calli were transferred into MSB liquid medium to establish suspension culture. When the suspended calli were transferred back onto MSB agar medium with 0. 1 mg /L IAA, 0. 5mg/L KT, 5% mannitol （cultured in light,2000 lx,12h/d）, a lot of adventitious roots formed in 7 — 10 days, and then somatic embryos formed from the protoplast derived calli. But o-nly a few embryoids developed further into the cotyledonary stage ,and the others died at globular, heart-shaped, or torpeto stage .Finally, some cotyledonary embryoids germinated and developed into plantlets or shoots with leaves.更多还原