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培养基中添加不同浓度的卡那霉素、头孢霉素和羧苄青霉素 ,观察抗生素对大白菜基因型GP 11种子发芽以及离体子叶再生的影响。结果表明 :GP 11种子在含有 2 0 0mg/L卡那霉素的MS培养基上发芽生长时 ,幼苗子叶完全黄化 ,侧根数为 0 ;随卡那霉素浓度的增加 ,芽、主根和侧根生长均受到一定程度抑制 ,但发芽率和发芽势不受影响。 4日苗龄的离体子叶在含有 10mg/L卡那霉素并附加一定激素的改良MS培养基生长时 ,基本无绿芽诱出 ,在含 7 5mg/L和 5mg/L卡那霉素的培养基中 ,诱出的绿芽率只有 32 5 %和 4 0 2 % ;在仅含 3mg/L卡那霉素的生根培养基中 ,小苗完全丧失生根能力。在添加浓度高至 4 0 0mg/L的头孢霉素或羧苄青霉素的诱芽培养基中 ,GP 11诱芽率为 0 ,且它们之间差异不明显 ,但在添加头孢霉素或羧苄青霉素的生根培养基中 ,小苗生根依然正常。试验结果表明 ,在农杆菌介导的基因转化中 ,对大白菜转化苗筛选的卡那霉素浓度在 10mg/L左右是适宜的 ,添加的抑菌剂浓度在能控制农杆菌生长的同时 ,尽量降低浓度 ,最好不超过 4 0 0mg/L ;在诱导生根培养基中 ,卡那霉素浓度不宜超过 3mg/L。另外 ,对T0 代转基因种子进行遗传分析时在发芽培养基中添加卡那霉素浓度应该达到2 0 0mg/L。
Different concentrations of kanamycin, cefotaxime and carbenicillin were added into the medium to observe the effect of antibiotics on the germination and regeneration of detached cotyledon of Chinese cabbage GP 11 seeds. The results showed that the seedlings cotyledons were completely yellowed on GP 11 seeds when germinated on MS medium containing 200 mg / L kanamycin. The number of lateral roots was 0; with the increase of kanamycin concentration, The growth of lateral roots was inhibited to a certain extent, but the germination rate and germination potential were not affected. The detached cotyledons of 4-day-old seedlings were almost free of green shoots when cultured in the modified MS medium containing 10 mg / L kanamycin and a certain amount of hormones. In the medium containing 75 mg / L and 5 mg / L kanamycin , Only 32.5% and 40.02% of green shoots were elicited. In rooting medium containing only 3 mg / L kanamycin, seedling completely lost rooting ability. In the induction medium of cefotaxime or carbenicillin with the highest concentration of 400 mg / L, the rate of bud induction of GP 11 was 0, and there was no significant difference between them. However, with the addition of cefotaxime or carbobenzyl Penicillin rooting medium, seedling rooting is still normal. The results showed that in the Agrobacterium-mediated gene transformation, Kanamycin screening of Chinese cabbage transformation of the concentration of 10mg / L or so is appropriate, the concentration of bacteriostat added can control the growth of Agrobacterium at the same time, Try to reduce the concentration, preferably not more than 400mg / L; in inducing rooting medium, kanamycin concentration should not exceed 3mg / L. In addition, the genetic analysis of T0 transgenic seeds in germination medium added kanamycin concentration should reach 200 mg / L.