| 猪繁殖与呼吸道综合征病毒荧光定量PCR检测方法的研究 |
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| 引用本文： | 郝贵增,付亮,张福良,李敬玺.猪繁殖与呼吸道综合征病毒荧光定量PCR检测方法的研究[J].安徽农业科学,2012,40(14):8099-8101. |
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| 作者姓名： | 郝贵增 付亮 张福良 李敬玺 |
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| 作者单位： | 安阳工学院生物与食品工程学院,河南安阳,455000;河南科技学院,河南新乡,453003 |
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| 基金项目： | 河南省科技攻关重点项目 |
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| 摘 要： | 通过条件优化,以定量的10倍系列稀释的质粒pMD-ORF7为标准品,进行荧光定量PCR扩增并制作标准曲线,建立了PRRSV的荧光定量RT-PCR检测方法。结果表明,该方法检测灵敏度可达1.0×100拷贝/μl;利用该方法对3份不同的组织样品进行重复性检测,计算出平均循环阈值的标准差分别为0.14、0.14和0.44,变异系数分别为0.01%、0.01%和0.02%,说明该方法具有良好的重复性和重现性。对阳性组织病料的检测表明,该方法与常规RT-PCR的阳性符合率为100%,但其检测灵敏度较常规RT-PCR约高100倍。
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| 关 键 词： | 猪繁殖与呼吸综合征病毒 荧光定量PCR Taqman探针 |
Establishment of Fluorescent Quantitative PCR Assay by TaqMan Probe Approach for PRRSV Detection |
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| Institution: | HAO Gui-zeng et al(School of Biotechnology and Food,Anyang Institute of Technology,Anyang,Henan 455000) |
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| Abstract: | The recombined plasmid pMD-ORF7 was constructed and used as standard positive template.And the FQ-PCR assay was constructed by quantitative concentration of serial 10 fold dilutions of pMD-ORF7 cDNA by optimizing circulation parameters.A standard curve was achieved.The result showed the sensitivity of this method was 1.0×100copy/μl.To detect three different samples by FQ-PCR repeatedly,the average circle threshold value was 0.14,0.14 and 0.44,with the coefficient of variation of 0.01%,0.01% and 0.02%,respectively.The FQ-PCR was proved to be with high sensitivity and repeatability.The detection of PRRSV in positive samples by both FQ-PCR and conventional RT-PCR was 100%,but the sensitivity of the former was 100 times higher than that of the latter. |
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| Keywords: | Porcine reproductive and respiratory syndrome virus Fluorescent quantitative PCR Taqman probe |
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