美洲型猪繁殖与呼吸综合征病毒TaqMan荧光RT-PCR检测方法的建立

Establishment of TaqMan real-time RT-PCR assays for detection of north american type porcine reproductive and respiratory syndrome virus

根据猪繁殖与呼吸综合征病毒(PRRSV)美洲株的基因序列,设计U1和U6两套PCR引物和荧光探针,建立了扩增U1和U6基因的荧光RT-PCR用于检测美洲型PRRSV。用10倍倍比稀释的质粒DNA、cDNA、RNA进行扩增以检测其灵敏度,同时对猪瘟(HCV)、猪伪狂犬(PRV)等7种病毒进行特异性检测。结果显示建立的扩增U1和U6基因的荧光RT-PCR可用于检测PRRSV,其灵敏度为10个拷贝的质粒DNA,而与HCV等非PRRSV无交叉反应。本研究所建立的荧光RT-PCR具有快速、灵敏、准确、低污染等优点,可用于PRRSV样品的检测。

Two pairs of real-time RT-PCR primers and probes, U1 and U6 were designed and synthesized based on the nucleotide sequences of the porcine reproductive and respiratory syndrome virus （PRRSV）. The minimal detection threshold of U1 and U6 real-time RT-PCR were approximately 10 copies of plasmid DNA, The specificity of the two methods was demonstrated by detecting 7 viruses including classical swine fever virus （HCV）, pseudorabies virus （PRV）. It was concluded that the real time RT-PCR could be effective methods for rapid detection of PRRSV.