禽偏肺病毒RT-PCR检测方法的建立及应用  

Establishment and application of a RT-PCR method for detection of avian metapneumovirus

作　　者：陈琳[1] 刁有祥[1] 邹金峰[1] 

机构地区：[1]山东农业大学动物科技学院,山东泰安271018

出　　处：《中国兽医学报》2012年第7期971-974,996,共5页Chinese Journal of Veterinary Science

基　　金：山东省科技发展计划资助项目(2010GNC10912)

摘　　要：根据GenBank中已经发表的B亚型禽偏肺病毒(aMPV)F基因的保守序列设计并合成1对引物,利用RT-PCR可以扩增出1条725bp的片段,进行特异性试验和敏感性试验,建立了禽偏肺病毒病的RT-PCR检测方法。特异性试验表明,建立的RT-PCR检测方法能够从禽偏肺病毒疫苗毒株VIR 115-B中扩增到725bp的特异性片段,而对H9N2亚型禽流感病毒、新城疫病毒、传染性支气管炎病毒的扩增结果均为阴性;敏感性试验表明,该方法最低检出量的cDNA质量浓度为1.45μg/L;对山东省492份病料进行检测,阳性检出率为43.09%(212/492),随机挑取11份进行克隆测序及序列分析,结果显示所扩增到的阳性产物均为B亚型的禽偏肺病毒。建立的禽偏肺病毒的RT-PCR检测方法具有快速、准确、特异性强、敏感性高的特点。A rapid and accurate detection method was established for avian metapneumovirus. According to the ge- nomic sequences of F gene of aMPV published in GenBank,a pair of primers was designed for amplifying the 725 bp fragment in RT-PCR experiments and followed by specificity test and sensitivity test. The specificity test showed that the RNA of aMPV could specially amplify the 725 bp fragment, but other nucleotide extracted from HgN2, NDV and IBV were negative. The sensitivity test showed that as low as 1.45 t^g/L of aMPV＇s cDNA could be de- tected by RT-PCR method. Then 492 samples of broilers were detected and the positive rate was 43.09 % （212/ 492）. We randomly picked 11 positive samples from different areas for further confirmation. Based on the analysis we could confirm that all the positive samples belonged to subtype B. The established RT-PCR diagnostic method is fast,accurate, highly sensitive and specific for the detecting avain metapneumovirus.

关 键 词：禽偏肺病毒 RT—PCR 检测 

分 类 号：S852.65[农业科学—基础兽医学]