农杆菌介导法获得桃叶卫矛转Bt基因植株  

Agrobacterium tumefaciens-mediated Transformation of Bt Gene in Euonymus bungeanus Maxim.

作　　者：张敏 杨承香 蒋子凡 马艳 Zhang Min;Yang Chengxiang;Jiang Zifan;Ma Yan(College of Horticulture,Jinling Institute of Technology,Nanjing,210000;Comprehensive Service Center of Jionggang Town,Dongtai,224237)

机构地区：[1]金陵科技学院园艺园林学院,南京210000 [2]弶港镇综合服务中心,东台224237

出　　处：《分子植物育种》2021年第19期6406-6412,共7页Molecular Plant Breeding

基　　金：江苏省林业科技创新与推广项目(LYKJ[2019]31);金陵科技学院高层次人才工作启动项目(JIT-B-202107)共同资助。

摘　　要：虫害一直是制约农林作物高产、优产和稳产的重要因素,开展植物抗虫性研究具有关键的理论和实践意义。本研究使用农杆菌介导法,以桃叶卫矛(Euonymus bungeanus Maxim.)的愈伤组织为外植体材料,研究农杆菌的菌液浓度、侵染时间及共培养时间对桃叶卫矛叶片愈伤组织转化效率所产生的影响;并在建立的最佳转化体系下将苏云金芽孢杆菌(Bacillus thuringiensis) Bt基因及豇豆胰蛋白酶抑制剂(Cowpea trypsin inhibitor) SCK基因导入桃叶卫矛中,最终获得稳定表达的转化植株。结果显示,在侵染菌液OD_(600)=0.5~0.6,侵染时间30 min,共培养5 d的条件下,桃叶卫矛叶片愈伤组织的转化效率最好,经PCR鉴定检测后,在23株抗性植株中有10株同时检测到两个抗虫基因的表达,证明外源抗虫基因已导入桃叶卫矛基因组中。本研究将为卫矛科植物的抗虫性育种工作提供一定的理论基础。Insect pests have always been an important factor limiting high, excellent and stable yields of agricultural and forestry crops, and it is of key theoretical and practical significance to conduct research on plant pest resistance.In this study, we used the Agrobacterium tumefaciens-mediated method to study the effects of Agrobacterium concentration, infiltration time and co-culture time on the transformation efficiency of Euonymus bungeanus Maxim.The Bt(Bacillus thuringiensis) gene and SCK(cowpea trypsin inhibitor) gene were introduced into E. bungeanus under the established transformation system, and the stably expressed transformed plants were finally obtained.The results showed that the best transformation efficiency of E. bungeanus leaf callus tissue was achieved under the conditions of OD_(600)=0.5~0.6, 30 min of infestation time and 5 d of co-cultivation. After PCR identification, the expression of two insect resistance genes was detected simultaneously in 10 of the 23 resistant plants, proving that exogenous insect resistance genes had been introduced into the E. bungeanus genome. This study will lay a foundation for the breeding work of insect resistance of Celastraceae.

关 键 词：桃叶卫矛 农杆菌介导法 胚性愈伤组织 遗传转化 

分 类 号：Q943.2[生物学—植物学] S332.3[农业科学—作物遗传育种]