西洋参叶肉原生质体的游离与培养

Primary Discussion on Leaf Protoplast Dissociation and Culture of America Ginseng

[目的]为了探讨西洋参叶肉原生质体游离和培养的最佳条件。[方法]以5年生西洋参为材料,研究了纤维素酶的种类,酶的浓度和渗透压对西洋参叶肉原生质体游离的影响,并探讨了西洋参叶肉原生质体在KM8P、MS基本培养基中的分裂情况。[结果]在混合酶液其他组分相同的情况下,日本产R-10纤维素酶的游离效果最好。酶浓度为4%时,3 h后大部分叶片都被解离成原生质体。幼嫩叶片的原生质体产量最高,为4.8×105个/g。KM8P培养基适合于西洋参叶肉原生质体的培养,较高的2,4-D浓度可以促进分裂。[结论]用混合酶系统处理,能够获得大量(4.8×105个/g)有活性的原生质体。游离原生质体的酶浓度以1%～2%为宜。KM8P+2,4-D(2.0 mg/L)+6-BA(2.0 mg/L)+KT(0.7 mg/L)培养基适宜培养西洋参叶肉原生质体。

[Objective] The study aimed to discuss the optimum conditions of leaf protoplast dissociation and culture of America ginseng. [Method] With 5 -year -old America ginseng as material, the effects of eellulase species, enzyme concentrations and osmotic pressure on leaf protoplast of America ginseng were studied and the dissociation of America ginseng leaf protoplast on KM8P and MS basic medium was discussed. [Result] When the other components of mixed enzyme liquid was same, the dissociation effect of R-10 eeIIulase made in Japan was best. When the enzyme concentration was 4%, most leaves were dissociated into protoplasts after 3 h. The protoplast output of young leaves was highest, being 4.8 × 10^5 per gram. KM8P medium was proper to cultivate America ginseng leaf protoplast and 2,4-D with higher concentration could promote the dissociation. [Conclusion] A lot of activated protoplasts （4.8× 10^5/g）were got through mixed enzyme system treatment. The enzyme concentration for dissociating protoplast should be 1%-2 % and medium for cultivating America ginseng leaf protoplast should be KM8P ＋ 2,4-D（2.0）＋ 6-BA（2.0）＋ KT（0.7）.