大豆悬浮细胞培养及作为外源基因转化受体的研究  

Culture and Exogenous Gene Transformation of Suspension Cells in Soybean

作　　者：方星[1] 胡倩倩[1] 王建国[1] 岳娜[1] 孟祥勋[1] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123

出　　处：《大豆科学》2015年第1期36-41,共6页Soybean Science

基　　金：吉林省科技发展计划重点项目(20060202)

摘　　要：以吉林小粒7号大豆品种为材料,切取无菌幼苗下胚轴诱导愈伤组织,初步建立了大豆悬浮细胞系,测定不同硼酸浓度下悬浮细胞生长变化;采用农杆菌介导法对大豆悬浮细胞进行外源GUS基因的转化,先通过不同程度的超声处理确定最佳超声时长,然后在最佳超声处理时长的基础上,探讨了硼酸浓度对转化效率的影响;最后采用PCR和Southern blot杂交对抗性愈伤做分子鉴定。结果表明：大豆悬浮细胞正常生长硼酸浓度在2.0-10 mg·L^-1,过高或过低均会抑制大豆细胞正常的生长,高于100 mg·L^-1时抑制作用最显著。但在转化过程中,适当增加硼酸浓度（10-30 mg·L^-1）则能够提高悬浮细胞的转化效率,并且有助于转化组织的筛选,借以3 min超声的辅助作用,30 mg·L^-1硼酸浓度下转化效率达到1 mg转化细胞团中含28个转化体。In the experiment,a soybean suspension cells was initially established with the callus induced from the hypocotyl of sterile seedling of Jilinxiaoli 7 soybean. And then the influence of concentrations of boric acid on the growth of suspension cell cultures（ fresh weight） were determinated and the transformation of exogenous GUS gene to the cells were conducted by using Agrobacterium-mediated method. The transformation efficiencies influenced by sonication duration and boric acid concentration were mainly investigated through histochemical method in order to find an appropriate genetic transformation way. The positive transformed cell cultures were further identified by PCR technique and Southern blot hybridization. The results showed that boric acid concentration for normal growth of soybean suspension cell was between 2. 0 and 10 mg · L^-1,higher or lower boron medium would inhibited soybean cell growth,especially at a high level of more than 100 mg · L^-1. However,the appropriate increase in boric acid concentration（ 10-30 mg · L^-1） could improve the transformation efficiency of the cell suspension,which was also helpful to the screening of the transformant in the transformation process. Twenty eight transformants were obtained in 1 mg transformed cells under 30 mg·L^-1boric acid concentration with the assistance of 3 min sonication.

关 键 词：大豆 悬浮细胞 硼酸 农杆菌 超声 转化 

分 类 号：S565.1[农业科学—作物学]