多重RT-PCR一步法技术同时检测猪瘟病毒和蓝耳病病毒方法的建立以及初步应用

猪瘟病毒和蓝耳病病毒均能导致猪繁殖障碍,对养猪生产影响很大。根据猪瘟病毒(CSFV)和猪蓝耳病(PRRS)的基因保守序列设计了2对针对这2种病毒的特异引物,并建立了多重RT-PCR方法,分别对其最佳反应条件、特异性及敏感性进行了测定,结果表明能同时扩增得到2条与试验设计相符的167bp(CSFV)和320bp(PRRS)特异性条带,同时具有较好的特异性;敏感性检测结果表明,临床阳性的样品提取的核酸稀释1000倍后仍能检测出CSFV和PRRSV。本方法的建立对于这2种病毒病的早期快速检测具有十分重要的意义。

Establishment and Primary Application of Multiplex Polymerase Chain Reaction for Detection of Classical Swine Fever and Porcine Reproductive and Respiratory Syndrome Virus

A multiplex polymerase chain reaction (M-PCR)was established and optimized to detect swine fever virus (CSFV)and porcine reproductive and respiratory syndrome virus (PRRSV)simultaneously. Two sets of specific primers were designed according to the conservative sequence of CSFV and PRRSV in the GenBank.It was showed that all sampies which contained CSFV or PRRSV could be amplified by the multiplex PCR using these two sets primers.Two specific bands of CSFV 167 bp and PRRSV 320 bp were detected using agarose gel electrophoresis in this multiplex PCR and accorded with designed result.It could detect 1 000 diluted nucleic acid of CSFV or PRRSV from clinic samples. The method provides a fast and reliable way of identifying CSFV or PRRSV. The method could prove very useful for laboratories working with early rapid identification of CSFV or PRRSV.