病毒诱导的基因沉默防控烟草马铃薯Y病毒病研究

  背景  病毒诱导的基因沉默（virus-induced gene silencing，VIGS）是植物抗病毒侵染的一种自然机制。  方法  以马铃薯Y病毒（Potato virus Y，PVY）脉坏死株系（PVYN）的外壳蛋白（coat protein, CP）基因为靶向序列，构建烟草脆裂病毒（Tobacco rattle virus，TRV）介导的基因沉默表达载体pTRV2-CP，转化农杆菌GV3101后，与pTRV1-GV3101混合成TRV: : CP（OD₆₀₀ 0.4~0.6）喷施处理剪叶后的烟苗，测定其侵染效率。人工接种PVY于对照及TRV: : CP处理的烟苗，RT-qPCR检测、表型观察和小区试验评价TRV介导的VIGS体系对PVY侵染的沉默效果。  结果  ① 与TRV: : 00相比，接种PVY 35 d时，TRV: : CP处理的烟株中PVY累积量减少了72.8%；②VIGS表达载体45 d内能有效表达；③温室盆栽和田间小区试验TRV: : CP处理对PVY防治效果分别达到85.12%和73.08%。  结论  研究建立的VIGS体系能够沉默PVY CP基因的表达，有效防治烟草马铃薯Y病毒病，为PVY的防治提供了新的方法。 

Control of tobacco Potato virus Y by virus-induced gene silencing

Virus-induced gene silencing (VIGS) is a natural mechanism for plant to resist viral infection. Using the coat protein (CP) gene of Potato virus Y (PVY) vein necrosis strain (PVYN) as the target sequence, the gene silencing expression vector pTRV2-CP mediated by Tobacco rattle virus (TRV) was constructed, transformed into Agrobacterium tumefaciens GV3101, and mixed with pTRV1-GV3101 to form TRV: : CP (OD₆₀₀ 0.4~0.6), which was then sprayed on tobacco seedlings after leaf cutting, so as to determine the infection efficiency of tobacco seedlings. Tobacco seedlings treated with TRV: : CP were inoculated with PVY artificially. RT-qPCR, phenotypic observation and plot test were used to evaluate the silencing effect of TRV-mediated VIGS system on PVY infection. It was showed that the infection rate of TRV: : CP was 95.24%. Compared with TRV: : 00, the cumulative quantity of PVY in tobacco plants treated with TRV: : CP decreased by 72.8% at 35 days after inoculation, and the expression vector of VIGS could be effectively expressed within 45 days. The control effects of TRV: : CP on PVY in greenhouse pot and field plot experiments were 85.12% and 73.08%, respectively. The VIGS system established in this study can silence the expression of PVY CP gene and effectively control tobacco PVY disease, which provides a new method for the prevention and control of PVY.