文献类型：期刊文献

中文题名：猴B病毒通用核酸检测体系的建立与评价

英文题名：ESTABLISHMENT AND EVALUATION OF A UNIVERSAL NUCLEIC ACID TEST METHEOD FOR DETECTING MONKEY B VIRUS

作者：马丽敏[1,2];贾俊婷[2];钟亚迪[2];范瑞[2];马玉媛[2];刘兴友[1,3];章金刚[2,3]

第一作者：马丽敏

机构：[1]河南科技学院;[2]军事科学院军事医学研究院卫生勤务与血液研究所;[3]新乡学院

第一机构：河南科技学院,新乡453000

年份：2018

卷号：26

期号：2

起止页码：22-27

中文期刊名：中国动物传染病学报

外文期刊名：Chinese Journal of Animal Infectious Diseases

收录：CSTPCD;;北大核心:【北大核心2017】；

基金：国家科技支撑计划项目(2015BAI07B00)

语种：中文

中文关键词：猴B病毒;实时荧光定量PCR;核酸检测

外文关键词：Monkey B virus;real-time fluorescent quantitative PCR;nucleic acid detection

摘要：建立能同时检测猴B病毒5种基因型的通用核酸检测体系,并对其进行方法学评价。首先,对猴B病毒5种基因型代表毒株全序列进行比对,选取其保守区域(g B基因)设计猴B病毒的通用引物和探针。其次,构建参考品质粒,并以其为模板建立核酸检测体系的标准曲线。最后,对该核酸检测体系分别进行特异性、通用性、敏感性和重复性评价。建立的标准曲线在3×107~3×102 copies模板范围内相关系数|r|=0.999。方法学评价结果显示,该核酸检测体系与人单纯疱疹病毒1型、人单纯疱疹病毒2型、水痘带状疱疹病毒、EB病毒、巨细胞病毒均无交叉反应;可同时检测猴B病毒5种基因型,检测下限为30 copies;批内和批间实验重复性良好。建立的猴B病毒通用核酸检测体系对于疑似猴B病毒感染病例的辅助诊断,以及高质量实验猴群的建立均有着重要意义。
To establish and evaluate a universal real-time fluorescent quantitative PCR method for identification and quantification of 5 Monkey B virus(macacine alphaherpesvirus 1)genotypes,a set of suitable universal primers and probes were designed from the conserved region of the gB gene available in the NCBI nucleotide database.Then,plasmids were constructed and used to generate standard curves.The quantitative PCR method was developed and assessed for its specificity,sensitivity and reproducibility.A linear relationship was observed from 3×102 copies to 3×107 copies of Monkey B virus.The qPCR developed here did not amplify Herpes simplex virus 1,Herpes simplex virus 2,Epstein-barr virus,Varicella zoster virus and Cytomegalovirus.The detection limit of qPCR assay was 30 copies for 5 genotypes of Monkey B virus.Analyses of the inter-and intra-assay variabilities showed good reproducibility.The establishment of the universal real-time qPCR method for the detection of Monkey B virus DNA would facilitate diagnosis of Monkey B virus infection in humans and make great significance to the generation of high-quality experimental monkeys.