甘蓝CMS下胚轴和子叶的离体培养与植株再生研究

Study on tissue culture and plant regeneration of hypocotyl and cotyledon of cytoplasmic male sterile line in cabbage

【目的】探索一种利用甘蓝胞质雄性不育系（CMS）下胚轴和子叶进行快速扩繁的技术，为利用CMS大量繁制甘蓝杂交种提供一条新途径。【方法】以甘蓝CMS05—2—10为试材，将苗龄5～7d无菌苗的下胚轴和子叶切下，接种于MS培养基上，45～50d后统计再生芽数，比较6一BA与NAA不同质量浓度配比对不定芽的诱导情况；将诱导出的不定芽接种于添加不同质量浓度（O，0．1，0．3，0．5mg／L）NAA的MS生根培养基上，20d后比较生根率和根的长势。【结果】在对下胚轴和子叶进行不定芽诱导时，MS培养基中分别添加1．0和4．0mg／L的6-BA对下胚轴、子叶的诱导效果较好，其芽再生频率和分化系数分别为83．3％，53．0％和3．6，3．0。在MS培养基中添加1．0mg／L6-BA＋0．2mg／LNAA，其对下胚轴不定芽的诱导效果较好，芽再生频率为80．7％，分化系数为3．4，褐化率为1．5％；在MS培养基中添加4．0mg／L6-BA＋0．3mg／LNAA，其对子叶不定芽的诱导效果较好，芽再生频率为50．0％，分化系数为3．2，褐化率为3．4％；二者的褐化率均明显低于仅添加6-BA的处理。在MS生根培养基中添加0．3mg／LNAA时，不定芽的生根效果较好，生根率达100％，且根系生长健壮。【结论】利用甘蓝胞质雄性不育系CMS05—2—10的下胚轴和子叶作为外植体进行不定芽的诱导，对于下胚轴，其适宜培养基为MS＋1．0mg／L6-BA＋30g／L蔗糖＋8g／L琼脂；对于子叶，其适宜培养基为MS-4-4．0mg／L6-BA＋30g／L蔗糖＋8g／L琼脂。不定芽诱导生根的最适培养基为MS＋0．3mg／LNAA＋30g／L蔗糖＋8g／L琼脂。

【Objective】 This experimentation aimed to explore fast propagation technology using hypocotyl and cotyledon of cytoplasmic male sterile line in cabbage to offer a way to produce hybrid cabbage by using cytoplasmic male sterile line.【Method】 Cytoplasmic male sterile CMS 05-2-10 of cabbage was studied.Hypocotyl and cotyledon taken from 5-7 d seedling were inoculated in MS medium.The number of adventitious buds after 45-50 days was counted,and the result of hormone combinations of 6-BA and NAA on adventitious buds of induction was compared;adventitious buds were inoculated on MS medium supplement with different concentrations（0,0.1,0.3,0.5 mg/L）of NAA.The condition of rooting rate and growth vigour of root was analyzed.【Result】 When adventitious bud of hypocotyl and cotyledon were induced,the effects of adding 1.0 and 4.0 mg/L 6BA in MS medium on adventitious bud regeneration were better,shoot regeneration frequency and coefficient of differentiation were 83.3%,53.0%and 3.6,3.0 respectively.The effect of adding 1.0 mg/L 6-BA＋0.2 mg/L NAA in MS medium on adventitious bud regeneration of hypocotyl was better,shoot regeneration frequency was 80.7%,coefficient of differentiation 3.4,browning rate 1.5%;The effect of adding 4.0 mg/L 6-BA＋0.3 mg/L NAA in MS medium on adventitious bud regeneration of cotyledon was better,shoot regeneration frequency was 50.0%,coefficient of differentiation 3.2,browning rate 3.4%.Both of them can significantly reduce browning rate in comparison with only adding 6BA.The effect of adding 0.3 mg/L NAA in MS medium on rooting culture was better,rooting rate of adventitious bud reached 100%,and the root can grow strongly.【Conclusion】 Taking the hypocotyl and cotyledon of cytoplasmic male sterile CMS 05-2-10 of cabbage as explants to induce adventitious bud,the suitable medium on adventitious bud regeneration of hypocotyl is MS＋1.0 mg/L6-BA＋30 g/L sucrose＋8 g/L agar;the suitable medium on adventitious bud regeneration of cotyledon is MS＋4.0 mg/L 6-BA＋30 g/L sucrose＋8 g/L agar.The suitable medium for root induction is MS＋0.3 mg/L NAA＋30 g/L sucrose＋8 g/L agar.