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实验构建了2个大豆贮藏蛋白Gy1基因种子特异性表达载体,并将其分别转入玉米愈伤组织中。载体1以潮霉素为筛选标记,将Gy1基因片段插入到含有种子特异启动子的植物表达载体pCAMBIA1301-7αP上,得到种子特异性表达载体pCAMBIA1301-7αP-Gy1;载体2以BADH为筛选标记,将Gy1基因插入到带有耐盐基因BADH的植物表达载体pCAMBIA1301-BADH上,得到安全性无抗生素选择标记的种子特异性表达载体pCAMBI-A1301-sbeⅡb-Gy1。通过农杆菌介导法将其分别导入玉米自交系H99的愈伤组织中,共获得132株再生植株;经过PCR检测,初步证明获得了34株转基因阳性植株。
Two soybean seed-specific expression vectors of Gy1 gene were constructed and transformed into maize callus respectively. The vector 1 was selected with hygromycin as the screening marker, and the Gy1 gene fragment was inserted into the plant expression vector pCAMBIA1301-7αP containing the seed-specific promoter to obtain the seed-specific expression vector pCAMBIA1301-7αP-Gy1. The vector 2 was labeled with BADH, The Gy1 gene was inserted into the plant expression vector pCAMBIA1301-BADH with salt tolerance gene BADH to obtain a seed-specific safety expression vector pCAMBI-A1301-sbeIIb-Gy1 without antibiotic selection marker. A total of 132 regenerated plants were obtained by introducing Agrobacterium tumefaciens into the callus of maize inbred line H99. 34 transgenic plants were initially proved by PCR.