农杆菌介导的小麦芽生长点和分蘖节遗传转化研究

Agrobacterium tumefactions-mediated Genetic Transformation of Apical Merited and Tillering Node in Bread Wheat

为了研究小麦芽生长点和分蘖节为受体的农杆菌转化的可行性,以小麦品种‘河农827’和‘河农58-3’为受体材料,以来自水稻的OsDREB为供体基因,研究农杆菌介导的小麦芽生长点和分蘖节转化体系。结果表明,2种方法均能获得转基因植株:芽生长点侵染法T0代的成活率低(30.02%),转化率高(PCR检测转化率达13.79%);分蘖节注射法T0代成活率高(100%),转化率低(PCR检出率为3.25%),综合比较2种方法的转基因效率,芽生长点侵染法为2.65%,分蘖节注射法为3.25%。对野生型小麦卡那霉素抗性筛选,确定了转基因小麦卡那霉素的筛选浓度为90mg/L较为合适。芽生长点侵染法共培养过程中的抑菌,加入羧苄青霉素的最佳浓度为300mg/L,有利于共培养后外植体有较低的染菌率。

In order to research the wheat apical merited and tillering node of Agrobacterium tumefactions-mediated transformation for the receptors of feasibility,genetic transformation of OsDREB gene from rice（Oryza sativa） into two wheat varieties mediated by Agrobacterium tumefactions was studied with wheat apical merited and tillering node as explants.The results showed that,both methods could obtain the PCR-positive transgenic plants：survival rate for T0 generation with apical merited was lower（30.02%） than that（100%） of T0 generation with tillering node injection,but transformation ratio（13.79） for the apical merited method was higher than that（3.25%） of tillering node method.The optimum concentration for kanamycin treatment was 90 mg/L by the wild-type kanamycin-resistance screening.The optimum carbenicillin was 300 mg/L to inhibit the growth of Agrobacterium tumefactions during the co-culture by bud growing point infection method,and it not only ensured low contaminated rate,but also improved bud regeneration rate.