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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。
Abstract：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 HSV1-infected cells,but only had a little effect on HSV1 infection,so taking gD as the target of siRNA against HSV1 needs further study.
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基本信息:
中图分类号:R346
引用信息:
[1]朱钦昌,任哲,张春龙,等.siRNA干扰HSV1 gD糖蛋白基因的研究[J].病毒学报,2007(01):22-27.
基金信息:
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)
2007-01-15
2007-01-15