摘要
依据非洲猪瘟病毒(ASFV)的vp73基因保守序列,设计了特异性引物与TaqMan探针,扩增片段69bp,建立了ASFV实时荧光PCR检测方法。与OIE推荐用于检测ASFV的实时荧光PCR和普通PCR方法进行比对,结果显示,本文建立的方法与OIE两种检测方法的特异性相当,灵敏度与OIE荧光PCR方法相当,但比普通PCR方法高出至少10倍。标准定量曲线显示,本文方法最低检测限可达10 copies/μL病毒DNA(相关系数为0.993046)。重复性试验的批内CV值为0.61%～1.14%,批间CV值为1.37%～3.14%,表明本文方法的重复性和稳定性良好。可见,本文建立的基于vp73基因的ASFV实时荧光PCR检测方法具备快速、准确、灵敏的特点,将为检验检疫部门防御ASFV传入我国提供更多技术支持。
A specific primer pair and a probe were designed according to vp73gene from ASFV with an amplicon of 69 bp,and the fluorescent real-time PCR method in detecting ASFV was founded.Compared with three OIE reference methods,the results showed that four methods showed similar specificity.But the developed method was as sensitive as OIE real-time PCR method,but at least 10times more sensitive than the other two normal PCR methods.A quantitative assessment of standard curve showed the LLD of the developed method was at least 10copies/μL of virus ge- nome equivalents（R=0.993 046）.Intra-assay and inter-assay CVvalues were 0.61%-1.14%and 1.37%-3.14% separately,which showed the method had good repeatability.With its high specificity,sensitivity and good repeat-ability,the developed method would become a valuable tool for the routine diagnostic procedures for ASFV.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2010年第9期1173-1178,共6页
Chinese Journal of Veterinary Science
基金
国家质检总局科研项目(No.2009IK003)
国家863课题(200710Z418)
关键词
非洲猪瘟病毒
实时荧光PCR
检测
African swine fever virus（ASFV）
fluorescent real-time PCR method
detection
