施马伦贝格病毒实时荧光RT-PCR检测方法的建立及应用  

Establishment and application of the real-time fluorescence RT-PCR for the Schmallenberg virus

作　　者：王建昌[1] 王金凤[1] 段永生[1] 张永宁[2] 李静[1] 

机构地区：[1]河北出入境检验检疫局检验检疫技术中心,河北石家庄050051 [2]中国检验检疫科学研究院,北京100029

出　　处：《中国兽医学报》2015年第11期1765-1769,共5页Chinese Journal of Veterinary Science

基　　金：国家质量监督检验检疫总局科研项目(2013IK054)

摘　　要：依据施马伦贝格病毒(Schmallenberg virus,SBV)S基因保守序列,设计了特异性引物和MGB荧光探针,建立了SBV实时荧光RT-PCR检测方法,构建标准曲线,进行特异性、敏感性和重复性等试验评价,并对绵羊和牛全血样品进行了检测。结果表明,所建立的SBV检测方法模板浓度与Ct值呈现良好的线性关系,相关系数R2为0.998;特异性强,仅对SBV RNA出现特异性扩增曲线;灵敏度高,对SBV RNA的检测灵敏度为10copies;重复性好,重复检测灵敏度CV<3%;对临床样品的检测均为SBV阴性。所建立的SBV实时荧光RT-PCR方法可以为SBV的诊断和流行病学调查提供一种快速、特异、敏感的检测手段。Schmallenberg disease emerged in the Europe since the summer and autumn in 2011,and is caused by the Schmallenberg virus（SBV）.The SBV infections mostly occurre in the ruminants,such as sheep,cattle and goat.In this paper,the specific primers and probe were designed based on the S gene of SBV,and the real-time fluorescence RT-PCR for SBV was established.Through the construction of standard curve,the specificity,sensitivity and reproducibility appraisal,the SBV template concentration showed the good correlation with the Ct values,and the correlation coefficient（R2）was 0.998;the method was highly specific for SBV,the detecting limit was10 copies for SBV RNA,and showed the good reproducibility with the Coefficient of variation（CV）less than 3%.and all the samples in the clinical sheep and cattle whole blood samples were negative for SBV.The real-time RT-PCR established in the paper was fast,specific and sensitive for the SBV detection and epidemiological investigation.

关 键 词：施马伦贝格病毒 S基因 MGB探针 

分 类 号：S852.65[农业科学—基础兽医学]