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建立了适用于多个大豆品种的萌动胚真空渗透辅助的外源基因转化方法,对影响农杆菌介导转化的有关参数进行了比较研究。确定的优化条件为:预培养3d,在菌种活化液的OD600值为0.6并加以200μmol.L-1乙酰丁香酮的农杆菌菌种活化液,侵染时间为6h,共培养3d。在优化条件下,将携带GUS基因的35S启动子驱动的表达载体pCAMBIA1304的根癌农杆菌菌株GV3101分别转入鲁豆11和潍6823中,获得510株鲁豆11再生植株和591株潍6823再生植株,其中,抗性再生植株分别为444株和462株。通过PCR和GUS检测,证明分别获得了13株和15株转基因植株,转化率分别为2.2%和2.5%。
A method of transformation of exogenous genes assisted by vacuum infiltration of embryo germinated for multiple soybean cultivars was established, and the relative parameters affecting the transformation mediated by Agrobacterium were compared. The optimized conditions were as follows: pre-incubation for 3 days, OD600 value of the strain activation solution was 0.6 and 200μmol.L-1 acetosyringone Agrobacterium strain activation solution, the infection time was 6h, coculture 3d. Under optimal conditions, Agrobacterium tumefaciens strain GV3101 carrying the 35S promoter driven expression vector pCAMBIA1304 of GUS gene was transferred to Luodou 11 and Weifu 6823, respectively, and 510 plants of Luodian 11 and 591 strains of Weixian 6823 were regenerated Among them, 444 and 462 resistant plants were regenerated respectively. By PCR and GUS detection, 13 and 15 transgenic plants were obtained, respectively, with conversion rates of 2.2% and 2.5%, respectively.