多重PCR检测猪细小病毒和猪伪狂犬病病毒的研究

Detection of porcine parvovirus and porcine pseudorabies virus by multiplex polymerase chain reaction

根据GenBank上已发表的猪细小病毒(Porcine parvovirus,PPV)的VP2基因序列和猪伪狂犬病病毒(Porcine pseudorabies virus,PRV)的gH基因序列,设计合成了两对特异引物,分别建立了PPV和PRV的单项PCR诊断方法,通过对扩增条件的筛选,最终成功地建立了PPV和PRV的复合PCR诊断方法,即利用一次PCR反应,可同时扩增PPV的751bp和PRV355bp的特异性片段,而扩增猪圆环病毒Ⅱ型(PCV-2)及相应的培养细胞(PK-15)核酸结果均为阴性,对PPV和PRV的最低检出量分别为100pg和10pg的DNA。该方法适合对PPV和PRV的联合检测和鉴别诊断。

Multiplex PCR was developed using two pairs of primers that were designed according to the published sequences of PPV and PRV, respectively. PCR for PPV and PRV were established under optimized conditions. Specific PCR products of 75 i bp for PPV and 355 bp for PRV could be simultaneously amplified in the multiplex PCR, whereas. No PCR products were detected from cultural cells challenged with PCV-2 and control cultural cells （PKoi5）, The method was capable of detecting the template DNA of 100 pg for PPV and 10 pg for PRV, and could effectively differentiate PPV, PRV in a mixed infection.