山羊痘病毒及口疮病毒ITR-059基因双重PCR检测方法的建立

发布时间：2018-03-25 14:50

本文选题：山羊痘病毒　切入点：口疮病毒　出处：《中国兽医科学》2017年05期

【摘要】：为实现山羊痘病毒(GTPV)和口疮病毒(ORFV)的同步快速检测,基于GTPV的ITR基因及ORFV的059基因,分别设计用于普通双重PCR的引物2对、用于二联实时荧光定量PCR的特异性引物及探针2套;探针分别用5′JOE-Eclipse3′及5′FAM-TAMRA3′荧光染料标记物进行标记。结果显示,所建立的GTPV-ITR-L-ORFV-059双重普通PCR方法,可实现对GTPV与ORFV的特异性双重检测,检测敏感度可达10~4copies/μL DNA;所建立的ITR-059二联探针实时荧光定量PCR方法可同时特异性检测ORFV和GTPV产生特异性荧光信号,二联探针的检测敏感度可分别达10~(2.87)和10~(2.52) copies/μL DNA。本研究初步建立了可同步特异性检测GTPV和ORFV的双重普通PCR及二联探针实时荧光定量PCR检测方法。
[Abstract]:In order to realize the synchronous and rapid detection of Goatpox virus (ITR) and aphthmic ulcer virus (ORF), two pairs of primers for common double PCR were designed based on ITR gene of GTPV and 059 gene of ORFV, and 2 sets of specific primers and probes for dual real-time fluorescent quantitative PCR were designed. The probes were labeled with 5 GTPV-ITR-L-ORFV-059 JOE-Eclipse 3 'and 5 FAM TAMRA3'fluorescent dyes respectively. The results showed that the established GTPV-ITR-L-ORFV-059 double ordinary PCR method could be used to detect the specificity of GTPV and ORFV. The sensitivity of detection is up to 10~4copies/ 渭 L DNA, and the real-time fluorescence quantitative PCR method with ITR-059 dual probe can detect the specific fluorescence signal of ORFV and GTPV at the same time. The detection sensitivity of the dual probe can reach 10 ~ (2. 87) and 10 ~ (2. 52)) copies/ 渭 L DNA respectively. In this study, a double ordinary PCR and a dual probe real-time fluorescence quantitative PCR method for the simultaneous detection of GTPV and ORFV were preliminarily established.
【作者单位】：云南省畜牧兽医科学院云南省热带亚热带动物病毒病重点实验室;
【基金】：云南省重点新产品开发计划项目(2014BB014) 云南省现代农业奶牛产业技术体系建设项目[云财农(2009)171号]
【分类号】：S852.654

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