Turner Anne-Marie W, Arbuckle Jesse H, Kristie Thomas M
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland.
Curr Protoc Microbiol. 2014 Nov 3;35:14E.5.1-27. doi: 10.1002/9780471729259.mc14e05s35.
Herpes Simplex Virus (HSV) is a human pathogen that establishes latency and undergoes periodic reactivation, resulting in chronic recurrent lytic infection. HSV lytic infection is characterized by an organized cascade of three gene classes; however, successful transcription and expression of the first, the immediate early class, is critical to the overall success of viral infection. This initial event of lytic infection is also highly dependent on host cell factors. This unit uses RNA interference and small molecule inhibitors to examine the role of host and viral proteins in HSV lytic infection. Methods detailing isolation of viral and host RNA and genomic DNA followed by quantitative real-time PCR allow characterization of impacts on viral transcription and replication, respectively. Western blots can be used to confirm quantitative PCR results. This combination of protocols represents a starting point for researchers interested in virus-host interactions during HSV lytic infection.
单纯疱疹病毒（HSV）是一种人类病原体，它会建立潜伏状态并周期性地重新激活，导致慢性复发性裂解感染。HSV裂解感染的特征是有组织地级联表达三类基因；然而，第一类即立即早期基因的成功转录和表达对于病毒感染的整体成功至关重要。裂解感染的这一初始事件也高度依赖宿主细胞因子。本单元使用RNA干扰和小分子抑制剂来研究宿主和病毒蛋白在HSV裂解感染中的作用。详细介绍病毒和宿主RNA及基因组DNA分离方法，随后进行定量实时PCR的方法，分别可以表征对病毒转录和复制的影响。蛋白质免疫印迹可用于确认定量PCR结果。这些实验方案的组合为对HSV裂解感染期间病毒-宿主相互作用感兴趣的研究人员提供了一个起点。
