寄主诱导的基因沉默对辣椒疫霉菌的影响

Host-induced Gene Silencing is Effective against Phytophthora capsici in Nicotiana benthamiana

由辣椒疫霉菌(Phytophthora capsici)引起的辣椒疫病是生产中最严重的病害之一。本研究利用病毒诱导的基因沉默(virus induced gene silencing, VIGS)技术,通过瞬时表达疫霉菌基因沉默信号,在本氏烟草(Nicotiana benthamiana)-疫霉菌互作系统中建立寄主诱导的基因沉默(host-induced gene silencing,HIGS)体系,为防治辣椒疫病提供新思路。首先,在TRV2-GFP和对照TRV2-GUS的本氏烟叶片上接种表达GFP的致病疫霉菌菌株14-3-GFP,结果显示TRV2-GFP植株中致病疫霉菌的GFP表达显著降低,表明利用VIGS技术构建的HIGS体系在本氏烟草-疫霉菌体系中可行。此外,选取辣椒疫霉菌致病相关基因 PcAvh1、 Pchmp1和 PcGK4,构建到TRV2载体上,通过在本氏烟草中表达结合接种辣椒疫霉菌,结果显示,与对照相比植物生长表型无明显变化,表达TRV2- PcGK4的植株对辣椒疫霉菌表现抗病,表达TRV2- PcAvh1和TRV2- Pchmp1的植株对辣椒疫霉菌的抗感性没有显著影响,表明 PcGK4可能是辣椒疫霉菌致病必须的基因之一。以上结果表明,利用HIGS技术沉默辣椒疫霉菌 PcGK4基因可有效降低辣椒疫霉菌对寄主的侵染,HIGS技术可用于辣椒疫霉菌致病关键基因的筛选和鉴定。

Phytophthora capsici is one of the most notorious diseases in pepper production. This study, we used VIGS technique to transient express gene silencing signals of Phytophthora to establish the Nicotiana benthamiana-Phytophthora host induced gene silencing(HIGS) system to provide new ideas for controlling. First of all, we inoculated GFP labeled Phytophthora infestans isolate 14-3-GFP on TRV-GFP and TRV-GUS expressed N. benthamiana leaves, and observed a clear reduction of GFP fluorescent in mycelium of 14-3-GFPon TRV-GFP plants than control, which indicate the HIGS is applicable in Nicotiana benthamiana-Phytophthora system. Furthermore, we cloned the P.capsici virulence related genes PcAvh1, Pchmp1 and PcGK4, and constructed them into TRV2 vector. After expressing them in N.benthamiana, we inoculated P.capsici and screened for plant resistance. Results showed that, TRV- PcAvh1, TRV- Pchmp1 and TRV- PcGK4 plants showed no difference in plant growth and morphology. TRV- PcGK4 plants are resistant to P.capsici compering to control, showed by reduced lesion area. The lesion areas and P.capsici biomass of TRV- Pchmp1 and TRV- PcAvh1 plants were comparable to control. PcGK4 may be one of the necessary genes for P.capsici pathogenicity. Taken together, our results indicate PcGK4 is a good target for HIGS and HIGS technique can be used to screen and identify key pathogenic genes of P.capsici.