重组酶聚合酶扩增检测法快速检测猪圆环病毒 3 型。
Recombinase polymerase amplification assay for rapid detection of porcine circovirus 3.
机构信息
Hebei Academy of Science and Technology for Inspection and Quarantine, Shijiazhuang 050051, China; Center of Inspection and Quarantine Technology, Hebei Entry-Exit Inspection and Quarantine Bureau, Shijiazhuang 050051, China.
Institute of Animal Quarantine, Chinese Academy of Inspection and Quarantine, Beijing 100176, China.
出版信息
Mol Cell Probes. 2017 Dec;36:58-61. doi: 10.1016/j.mcp.2017.09.001. Epub 2017 Sep 25.
The objective of this study was to develop a real-time recombinase polymerase amplification (rt-RPA) assay for the rapid detection of porcine circovirus 3 (PCV3). Specific RPA primers and exo probes were designed for the cap gene of PCV3 within the conserved region of viral genome. The amplification was performed at 38 °C for 20 min. The rt-RPA was specific for PCV3, as there was no cross-reaction with other pathogens tested. Using the recombinant plasmid pUC57-PCV3 as template, the analytical sensitivity was 23 copies. Of the 186 clinical samples, PCV3 DNA was identified in the 51 samples by the rt-RPA, and the positive rate was 27.4% (51/186). The diagnostic agreement between the rt-RPA and real-time PCR was 96.2%. The R value of rt-RPA and real-time PCR was 0.919 by linear regression analysis. The developed rt-RPA assay shows promise for rapid and sensitive detection of PCV3 in diagnostic laboratories and at point-of-need, thus facilitating the prevention and control of PCV3.
本研究旨在开发一种用于快速检测猪圆环病毒 3（PCV3）的实时重组酶聚合酶扩增（rt-RPA）检测方法。针对 PCV3 基因组保守区的 cap 基因设计了特异性的 RPA 引物和外切探针。扩增在 38°C 下进行 20 分钟。rt-RPA 特异性检测 PCV3，与检测的其他病原体无交叉反应。使用重组质粒 pUC57-PCV3 作为模板，分析灵敏度为 23 拷贝。在 186 份临床样本中，通过 rt-RPA 在 51 份样本中鉴定出 PCV3 DNA，阳性率为 27.4%（51/186）。rt-RPA 和实时 PCR 之间的诊断一致性为 96.2%。线性回归分析显示，rt-RPA 和实时 PCR 的 R 值为 0.919。开发的 rt-RPA 检测方法有望在诊断实验室和现场快速灵敏地检测 PCV3，从而有助于 PCV3 的预防和控制。
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