【摘要】 为了能快速鉴别猪伪狂犬病病毒流行株，根据猪伪狂犬病病毒gE基因前半段95 bp～210 bp区域处设计一对特异性引物及探针，优化反应条件后，建立一种可鉴别猪伪狂犬病病毒gE缺失疫苗株与流行株的Taqman荧光定量PCR检测方法。结果显示，该方法对pMD-PRV gE阳性标准质粒的最低检出下限为4.4×10~1拷贝/μL；该方法特异性强，对猪常见病毒性疾病（猪瘟病毒、猪繁殖与呼吸综合征病毒、猪圆环病毒2型等）均无交叉反应，将PRV流行株和疫苗株（Bartha-K61株、HB2000株、C株、Ea株）同时与某商品化试剂盒对比检测，特异性优于某商品化荧光定量PCR检测试剂盒；组内和组间重复性试验的变异系数均小于1.6%，重复性好。应用本研究建立的荧光定量PCR检测方法对165份临床样品进行检测，阳性率为2.42%(4/165)，选取已知3份阳性的30份临床样品，与商品化检测试剂盒比对，符合率100%。因此，本研究建立的PRV gE基因荧光定量PCR检测方法可用于猪伪狂犬病病毒流行株的快速诊断和流行病学调查，为PR的控制与净化提供技术支持。更多还原

【Abstract】 In order to rapidly identify the epidemic strains of porcine pseudorabies virus, one pair of specific primers and probes were designed according to the region of 95 bp to 210 bp in the first half of the gE gene of porcine pseudorabies virus, and the reaction conditions were optimized to establish a Taqman fluorescence quantitative PCR assay that could identify the vaccine strain and the epidemic strains of porcine pseudorabies virus with g E deletion. The results showed that the lowest lower limit of detection of this method for pMD-PRV g E-positive standard plasmid was 4.4×10~1copies/μL; the method was specific, with no cross-reactivity to common viral diseases in pigs(swine fever virus, porcine reproductive and respiratory syndrome virus, porcine circovirus type 2, etc.), and the specificity of PRV epidemic strains and vaccine strains(Bartha-K61, HB2000, C, Ea strains) was better than that of a commercial fluorescent PCR kit; the coefficients of variation of intra-and inter-group reproducibility tests were less than 1.6%, which was good reproducibility. Applying the fluorescent quantitative PCR assay established in this study to 165 clinical samples, the positive rate was 2.42%(4/165), and 30 clinical samples with three known positives were selected and compared with the commercialized detection kit,with a 100% compliance rate. Therefore, the fluorescence quantitative PCR of PRV gE gene established in this study can be used for rapid diagnosis and epidemiological investigation of epidemic strains of porcine pseudorabies virus, and provide technical support for the control and purification of PR.更多还原