利用gfp报告基因优化农杆菌介导的水稻遗传转化

为提高农杆菌介导的水稻遗传转化效率,以晚粳97为转化材料,绿色荧光蛋白gfp基因为报告基因,采用正交试验L9(33)对影响农杆菌介导水稻的遗传转化因子进行优化。通过观察愈伤组织荧光表达情况,分析菌液浓度、共培养温度与共培养时间对农杆菌转化水稻的影响。结果表明,在OD660值为0.1、共培养21℃～23℃黑暗条件下,农杆菌与水稻愈伤共培养72 h,最有利于水稻的遗传转化,该条件下晚粳97愈伤组织荧光表达率达到70.9%。

Improvement of Agrobacterium-mediated genetic transformation of rice using gfp as a reporter gene

To improve the Agrobacteriam-mediated transformation frequency of Japonica rice Wanjing 97, an orthogonal experiment L9 （33） was designed to optimize the factors regulating rice genetic transformation, and the impacts of innoculum OD6oo, coculture temperature and coculture duration on transformation using gfp gene to monitor the expression of target gene were analyzed. Results showed that the optimal conditions for Agrobacterium-mediated transformation of Wanjing 97 were determined to be innoculum OD660 of 0. 1, cocuhured at 21℃ -23℃ dark for 72 h. Under this condition, the rate of gfp-positive calli was up to 70. 9%. The results suggested that gfp could be used as an effective reporter gene to improve rice genetic transformation.