靶向H、N基因siRNA对犬瘟热病毒在vero细胞中复制的抑制作用  

Inhibiting the replication of canine distemper virus(CDV) in Vero cells by siRNA targeting the nucleocapsid protein gene and the hemagglutinin protein gene

作　　者：袁龙[1] 艾纯旭[1] 马云青[1] 陈健[1] 袁宝[1] 高巍[1] 胡进平[1] 任文陟[1] 

机构地区：[1]吉林大学动物科学学院,吉林长春130062

出　　处：《中国兽医学报》2015年第7期1046-1050,共5页Chinese Journal of Veterinary Science

基　　金：吉林省科技发展计划青年科研基金资助项目(201201014);吉林大学基本科研业务费项目(450060481144);吉林大学农学部引进人才科研启动基金资助项目(4305050102Q7)

摘　　要：以犬瘟热病毒(CDV)的核衣壳蛋白基因(N基因)和血凝素蛋白基因(H基因)作为靶基因,各设计并体外合成小干扰RNA,分别为N1、N2、N3和H1、H2、H3。将6对siRNA分别转染非洲绿猴肾细胞(Vero)并在转染后接种CDV,48h后通过细胞病变(CPE)、间接免疫荧光、TCID50、荧光定量PCR等方法对2组6个siRNA抑制CDV在Vero细胞中增殖的效果进行比较研究。结果显示,转染细胞感染CDV 48h后,6个siRNAs干扰组细胞出现的绿色荧光明显少于对照组,病毒感染滴度分别是对照组的1/293、1/11、1/29、1/83、1/302和1/90,6个干扰组的CDV基因拷贝数分别是对照组的12.96%、57.75%、23.47%、31.82%、12.82%、15.58%。结果表明,6对siRNAs均对CDV在Vero细胞中的复制具有不同程度的抑制作用,且以H2组抑制效率最高。Six pairs of RNAs（siRNA）,named N1,N2,N3 and H1,H2,H3 and targeting nucleocapsid protein gene（N gene）and the hemagglutinin protein gene（H gene）respectively,were prepared by in vitro transcription.Then,each of them was transfected with vero cells from an African green monkey before infected with CDV.AT 48 hours post-infection,these infected cells were evaluated for their antiviral activities against the CDV strain through cytopathic effect（CPE）,indirect immuno-fluorescence microscopy（IFA）,TCID50,and realtime fluorescence quantitative PCR.Results demonstrated that there was obviously less green fluorescent in the interfered siRNAs,compared with the control groups.The virus titers in the comparisons between the interfered groups and the control groups were 1/293,1/11,1/29,1/83,1/302 and 1/90,respectively.The number of CDV genes copied in the infected groups was 12.96%,57.75%,23.47%,31.82%,12.82% and 15.58% of the control groups,respectively.As is seen from the above,the six pairs of siRNAs had different effects on inhibiting the copy of CDV in Vero cells,among which H2 had the best effect.

关 键 词：靶向H、N基因 SIRNA 复制 

分 类 号：S852.65[农业科学—基础兽医学]