摘要： 为探讨RNAi策略在抗GFLV基因工程中的效果,通过RT-PCR克隆获得了310 bp的GFLV外壳蛋白基因保守片段,采用Gateway技术将该基因片段连入目标载体pHELLSGATE12构建了RNAi植物表达载体,并通过农杆菌介导法转入烟草中,RT-PCR检测表明目的基因片段已整合到烟草基因组中并在RNA水平上得到表达,病毒接种结果显示阳性植株的发病率明显低于对照,说明在转基因植株中dsRNA表达可干涉GFLV侵染过程。

关键词: 葡萄扇叶病毒, RNAi, 载体构建, 转基因, 烟草

Abstract: In order to probe the efficiency of the RNAi strategy in protection plants against GFLV,we cloned a conserved segment of coat protein gene of GFLV (310 bp) by RT-PCR,constructed a RNAi plant expression vector by introducing this fragment into the destination vector of pHELLSGATE12 by gateway technology,and transformed into Nicotiana benthamiana by Agrobaterium mediated method.RT-PCR identification showed that the target gene fragment was integrated into the tobacco genome and showed expression in RNA level.We also evaluated the virus-resistance of the plants by virus inoculation.The results showed that the rate of disease occurrence in the transgenic positive lines was obviously lower than the control,indicating that the expressed dsRNAs in the transgenic plants may interfere the infection of this virus.

Key words: Grapevine fanleaf virus, RNA interference, Vector construction, Transgenosis, Nicotiana benthamiana

中图分类号: 

• Q78