双重荧光定量RT-PCR快速检测肠道病毒和水痘-带状疱疹病毒  

Rapid detection of enterovirus and varicella-Zoster virus by multiplex real-time quantitative RT-PCR

作　　者：黄世旺[1] 李剑[1] 卢亦愚[2] 严菊英[2] 方叶珍[1] 倪志敏[1] 

机构地区：[1]杭州市江干区疾病预防控制中心,浙江杭州310004 [2]浙江省疾病预防控制中心,浙江杭州310051

出　　处：《中国卫生检验杂志》2013年第10期2274-2276,共3页Chinese Journal of Health Laboratory Technology

摘　　要：目的建立双重荧光定量RT-PCR技术,用于同时快速检测肠道病毒(Enteroviruses,EV)和水痘-带状疱疹病毒(Varicella-Zoster virus,VZV)。方法从GenBank上下载不同年份和地区的EV和VZV基因序列,分别在其保守区域设计引物与TaqMan探针,建立优化单重和双重荧光定量RT-PCR反应体系,评价所建双重RT-PCR反应体系的特异度、敏感度和稳定性,并应用于临床标本的检测中。结果该方法对EV和VZV检测具有高度特异性,检出限分别为30 copies/μl和36copies/μl,从疑似EV/VZV患者临床标本中直接检测EV与VZV,整个操作过程仅需3 h。结论本研究建立的双重荧光定量RT-PCR方法可以同时鉴别EV与VZV,具有特异性好、灵敏度高、快速易操作等优点,适用于EV/VZV早期感染病例的快速确认以及暴发疫情的应急诊断。Objective To establish a TaqMan - based multiplex real - time RT - PCR assay for detection of entero- viruses and Varicella - Zoster virus. Methods The gene sequences of EV and VZV downloaded from the Genbank database were aligned using the biologic software, and the specific primers and probe were designed in the 5＇ un- translated conserved region for Ev and the conserved region of the ORF62 gene for VZV. The reaction conditions were optimized to evaluate the sensitivity, specificity and stability, and clinical specimens were detected by this as- say. Results This assay had high specificity for detection of EV and VZV, and the detection limits were 30 cop- ies/μl and 36 copies/μl. This assay can be directly used to identify EV and VZV from clinical specimens, only 3h. Conclusion This assay is rapid, sensitive and specific for early clinical detection and emergent examination of EV and VZV.

关 键 词：双重荧光定量RT-PCR TAQMAN探针 肠道病毒 水痘-带状疱疹病毒 

分 类 号：R446.5[医药卫生—诊断学]