根癌农杆菌介导的大豆子叶节转化体系的优化

以13种不同基因型的大豆为材料,研究了大豆外植体再生过程中不同大豆基因型对丛生芽数目的影响,选取优势基因型为受体材料,比较了在草丁膦和潮霉素筛选压力下外植体的再生情况,并确定其合适的筛选浓度,在对农杆菌介导的子叶节转化体系优化的基础上进行了大豆转化效率的研究。结果显示:在相同的芽诱导条件下,山宁14产生的丛生芽最多,更适合于大豆子叶节转化;与其它筛选剂相比,采用潮霉素的梯度筛选更有利于抗性苗的成活,其最适筛选浓度为8 mg.L-1;通过检测GUS基因的表达和分子鉴定证明外源的GUS基因已插入到转基因植株的基因组中,转化效率达到3.2%。

Optimization of Agrobacterium-mediated Soybean Transformation using the Cotyledonary Node

To optimize the system of Agrobacterium-mediated soybeanGlycine max(L.) Merrill]transformation,effects of 13 cultivars and 2 selection agents were studied during the process of shoot regeneration and transformation.The results showed that the cultivar ’Shanning 14’ induced the highest number of multiple shoot among tested 13 cultivars,and hygromycin B was better than glufosinate during transformant selection of cultivar ’Shanning 14’.The optimal transformant selection scheme was a gradient concentration of hygromycin B from 3 to 8 mg·L-1.The integrations of UidA(GUS)gene into the transformant genome were demonstrated by histological staining,PCR application and sequencing analysis.Transformation efficiency was 3.2% based on the number of transformed events.