多重PCR诊断猪细小病毒、伪狂犬病毒和圆环病毒Ⅱ型方法的建立和应用  

Multiplex PCR Assay for Detection of Porcine Parvovirus,Pseudorabies Virus and Porcine Circovirus Type

作　　者：刘洋[1,2] 李俊[3,2] 时建立[3,2] 孙文博[3,2] 徐绍建[3,2] 吴家强[3,2] 李坤[1,2] 于江[1,2] 王金宝[3,2] 周顺[1] 

机构地区：[1]青岛农业大学,山东青岛266109 [2]山东省畜禽疫病防治与繁育重点实验室,山东济南250100 [3]山东省农业科学院,山东济南250100

出　　处：《家畜生态学报》2010年第5期95-98,109,共5页Journal of Domestic Animal Ecology

基　　金：山东省自然科学基金项目(Y2006D23);兽医生物技术国家重点实验室开放基金课题(SKLVBF201007);十一五国家支撑计划(2006BAD06A18)

摘　　要：根据GenBank中已发表的猪细小病毒(PPV)、猪伪狂犬病毒(PRV)和猪圆环病毒Ⅱ型(PCV2)等3种病毒基因序列,对各病毒基因区进行同源性分析,确定PPV VP2、PRVgB、PCV2ORF1基因的保守区为各病毒的诊断靶序列。在建立各病毒单重PCR技术的基础上,优化多重PCR反应条件,建立了三种病毒的多重PCR技术。用78份临床病料对本研究多重PCR技术和单重PCR技术进行对比验证,结果显示,总符合率为97.4%以上。该方法特异性强、敏感性高,为PPV、PRV和PCV2临床诊断和流行病学调查等研究提供了新手段。Based on the similarity of the viral sequences deposited in the GenBank database,the VP2 gene of PPV,the gB gene of PRV,and the ORF1 gene of PCV2 were selected as the diagnostic targets.By using three pairs of virus-specific primers,three PCR assay were established to amplify the conservative regions of the three viruses,respectively.Consequently,a multiplex PCR method to detect the three viruses in one tube was developed.The multiple PCR system would amplify a 203 bp segments for PPV,a 368 bp segments for PRV and a 703 bp segments for PCV2 simultaneously or separately in the samples,depending on its infection status.To evaluate the multiplex PCR assay,78 clinical samples were comparatively detected.The data showed that the multiple PCR method being 97.4 coincidence with the single PCR.The results indicated that the optimized multiplex PCR is both sensitive and specific,providing a new method for the clinical diagnosis and epidemiological research of these three diseases.

关 键 词：多重PCR PPV PRV PCV2 

分 类 号：S811.5[农业科学—畜牧学]