新疆阿克苏地区猪圆环病毒3型TaqMan荧光定量PCR检测方法的建立

Establishment of fluorescence quantitative PCR for detection of porcine circovirus type 3 TaqMan in Akesu,Xinjiang

本研究在猪圆环病毒3型(PCV3)Cap基因保守区设计一对特异性引物和荧光探针,以PCV3 Cap基因阳性质粒为模板,建立PCV3的TaqMan荧光定量PCR检测方法。TaqMan荧光定量PCR检测标准曲线在3.72×10^(8)~3.72×10^(1)拷贝/μL之间有较好的结果。重复性试验结果中,组内变异系数在0.165%~0.588%之间,组间变异系数在0.013%~0.097%之间,变异系数均小于1%,表明本试验离散程度较小。在PCV3 TaqMan荧光定量PCR灵敏性试验中,灵敏性为3.72×10^(3)拷贝/μL,相较于传统PCR方法灵敏性增加10倍,本试验建立的TaqMan荧光定量PCR检测方法为快速精准检测PCV3奠定了基础。

A pair of specific primers and fluorescent probes were designed in the conserved region of porcine circovirus type 3(PCV3)Cap gene,and TaqMan PCR method for detection of PCV3 was established using the positive plasmid of PCV3 Cap gene as template.The standard curves of TaqMan PCR detection ranged from 3.72×10^(8) copies/μL to 3.72×10^(1) copies/μL could obtain good results.Repeatability tests showed that the intra-group and inter-group coefficients of variation range from 0.165%to 0.588%and 0.013%-0.097%,respectively,which were less than 1%,indicating that the dispersion degree of this test is small.In the sensitivity test of PCV3 real-time quantitative PCR detecting system,the sensitivity was 3.72×10^(3) copies/μL,which was 10 times than that of the traditional PCR method.The TaqMan PCR detection method established in this study laid a foundation for the rapid and accurate detection of PCV3.