农杆菌介导小麦遗传转化影响因素的研究.PDFVIP

河南农业科学, 2011,40(2) :4144 Journal of Henan Agricultural Sciences 贺 杰, 胡海燕, 周 岩, 赵俊杰, 魏琦超 ( , 453003) : 选用河南科技学院近年来选育的小麦品种百农160 的成熟胚愈伤组织和农杆菌菌株 LBA4404, 对农杆菌 导遗传转化中农杆菌浓度感染时间乙酰丁香酮浓度头孢霉素质量浓度 等几个重要影响因素进行了研究, 建立小麦高效转化体系结果表明, 菌液O 600= 0.6, 侵染时间 30min 时对愈伤组织的生存和转化最有利; 浓度为200mol/ L 的乙酰丁香酮可显著提高抗性愈伤 再生率; 随着头孢霉素浓度的升高, 愈伤组织出现褐化,生长缓慢, 再生率低,所以在有效抑制农杆 菌的浓度范围内,尽量采取较低的质量浓度在后期的抑菌分化筛选中,在继代培养基中添加头孢 霉素的质量浓度为500mg/ L, 继代扩增筛选培养2 个月, 每10~ 15d 换一次培养基; 而在分化培养 基中添加200mg/ L 头孢霉素,即可获得再生率高的抗性愈伤最适条件下转化频率达2.36 % : 小麦; 农杆菌转化; 侵染时间; 头孢霉素 : S512. 1 : A :2011 Factors Affecting Wheat Transformation Mediated by A g r obact er i um tumef aciens HE Jie, HU Haiyan, ZHOU Yan,ZHAO Junjie,WEI Qichao ( Henan Institute of Science and Technology,Xinxiang 453003, China) Abstract: To establish highly efficient wheat transformation system, factors which influenced wheat transformation including agrobacterium concentration, infection period, and concentration of acetosyringone and cephalosporin were studied with the mature embryo callus of wheat variety Bainong 160 and agrobacterium strain LBA4404. The results showed that the transformation effi ciency was clearly improved with agrobacterium density of O 600 = 0.6 and incubation for 30min. Acetosyringone with the concentration of 200mol/ L could significantly enhance the rate of re sistant calluses. Considering the inhibition of cephalosporin on agrobacterium cells, it was sug gested that 500mg/ L of cephalosporin should be used in the latter stage of antibacterial screen ing, and then 200mg/ L of cephalosporin should be used in differentiation medium to get resistant calluses of high regeneration efficiency. T he transgenic frequency was 2. 36% under the optimal conditions. Key words: Whea