猪传染性胃肠炎病毒与猪呼吸道冠状病毒荧光RT-PCR鉴别检测方法建立与应用

Establishment and Application of a TaqMan Real-time RT-PCR Assay for the Detection of Transmissible Gastroenteritis Virus and Porcine Respiratory Coronavirus

本研究根据Genbank登录的猪传染性胃肠炎病毒(TGEV)N基因和S基因保守区序列设计合成了两对引物和探针,通过对荧光定量RT-PCR反应条件的优化,建立了TaqMan荧光定量RT-PCR快速检测TGEV的方法。该方法能有效地鉴别序列密切相关的猪传染性胃肠炎病毒与呼吸道冠状病毒。与常规RT-PCR试验比较表明,所建立的荧光RT-PCR检测技术快速、敏感,检测时限3个小时以内,具有很好的特异性和重复性。通过对39份临床样品进行检测,结果表明所建立的检测方法直接检测样品中猪传染性胃肠炎病毒。

The probes and primers were designed and synthesized based on the conserved gene N and S of transmissible gastroenteritis virus （TGEV） available in GenBank, and the reaction parameters were optimized to develop a sensitive TaqMan-based real-time RT-PCR assay for the rapid detection of TGEV. The assay could differentiate TGEV from porcine respiratory coronavirus （PRCV） effectively. Compared with normal RT-PCR, the developed RT-PCR was fast and sensitive, with the detection time less than 3 hours. And the specificity and reproducibility of the assay were very good. By detecting 39 clinical samples, the results suggested that the method could be used to detect TGEV in samples effectively.