siRNA干扰HSV1 gD糖蛋白基因的研究  

Silencing HSV1 gD Expression in Cultured Cells by RNA Interference

作　　者：朱钦昌[1] 任哲[1] 张春龙[1] 张美英[1] 廖红娟[1] 刘秋英[1] 张佩琢 李久香[3] 胡巢凤[3] 王华东[3] 王一飞[1] 

机构地区：[1]暨南大学生物医药研究与开发基地,广东广州510632 [2]上海吉玛制药技术有限公司,上海201203 [3]暨南大学医学院,广州510632

出　　处：《病毒学报》2007年第1期22-27,共6页Chinese Journal of Virology

基　　金：广东省重大攻关项目(2005A10904001);广东省自然科学基金团队项目子课题(2004E039213);昆布海藻抗病毒作用研究及其功能食品的开发(2005B50301017);广东省生物工程药物重点实验室建设项目(2002B60119)

摘　　要：以HSV1 gD糖蛋白基因为靶点,设计合成5对siRNA,并构建pEGFP-N1-gD融合表达质粒,脂质体法共转染Vero细胞,利用绿色荧光蛋白(EGFP)报告基因的特征,流式细胞仪筛选特异沉默gD表达的siRNA。然后有效siRNA转染Vero细胞并感染HSV1,通过空斑减数实验,Real-time PCR和子代病毒滴度评价其对HSV-1感染的作用。结果显示siRNA能有效抑制gD-EGFP融合蛋白和感染细胞内gD糖蛋白的表达,但对HSV-1的感染性无明显抑制作用,故选择gD作为siRNA抗病毒靶点还需进一步的论证。To explore the anti-HSV-1 effect of silencing gD gene expression by RNA interference, five 21-nucleotide duplex small interfering RNAs（siRNAs） targeting the HSV1 gD sequence were designed and the gD-EGFP fusion gene expression vector was constructed, then co-tansfected into Vero cell, and screened the effective siRNA through analyzing the intensity of the EGFP fluorescence. Finally, the anti-HSV1 effect was confirmed by plaque reduction assay, real-time PCR and daughter virus titration of HSV1 infected Vero cells transfected with siRNAs. The study demonstrated that siRNAs could effectively and specifically inhibit gD gene expression in HSVl-infected cells, but only had a little effect on HSV1 infection, so taking gD as the target of siRNA against HSV1 needs further study.

关 键 词：单纯疱疹病毒1型 糖蛋白D RNA干扰 抗病毒 

分 类 号：R373.9[医药卫生—病原生物学]