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中国农学通报

中国农学通报 ›› 2011, Vol. 27 ›› Issue (3): 342-346.

所属专题： 畜牧兽医

• 畜牧 动物医学 蚕 蜂 • 上一篇    下一篇

鸡传染性支气管炎病毒RT-PCR快速检测方法的建立

刘金朋 商艳红 陈红英 王淑娟 王子馨 王亚丹 师丽刚   

  • 收稿日期:2010-10-11 修回日期:2010-11-22 出版日期:2011-02-05 发布日期:2011-02-05
  • 基金资助:

    国家“十一五”科技支撑计划专项

Development of RT-PCR for Rapid Detection of Infectious Bronchitis Virus

  • Received:2010-10-11 Revised:2010-11-22 Online:2011-02-05 Published:2011-02-05

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摘要：

根据鸡传染性支气管炎病毒(IBV) N基因序列（FJ767920），利用引物软件Premier5.0，设计并合成了1对特异引物，通过对RT-PCR扩增条件的优化，建立了一种快速检测IBV的RT-PCR方法。应用该方法成功地从IBV M41和河南株HN99中扩增出318 bp的目的片段，而传染性喉气管炎病毒、新城疫病毒及马立克病毒基因组均未扩增为出相应的片段。将回收的RT-PCR产物克隆到pGEM-T Easy载体后测序，进一步证实RT-PCR检测方法的特异性。对IBV的最低检出量为0.6 pg的cDNA。经临床初步应用表明，本方法的建立对IBV的早期诊断和临床检测具有快速、灵敏、特异、经济的特点。

关键词: 解读与传承, 解读与传承

Abstract:

A pair of primers was designed according to a conserved region of infectious bronchitis virus (IBV) N gene （FJ767920）, reverse transcriptase-polymerase chain reaction (RT-PCR) for rapid detection of IBV was developed by optimizing amplification conditions such as concentrations of primers for RT-PCR. A 318 bp fragment was amplified from IBV M41 strain and Nephropathogenic IBV HN99 strain, respectively, but not from infectious laryngotracheitis virus, Newcastle disease virus and Mareks disease virus. This amplified fragment was cloned into pGEM T-Easy vector and sequenced. The result showed this amplified fragment is specific for IBV cDNA. This method could detect the template cDNA of 0.6 pg for IBV. Clinical detection of IBV by RT-PCR showed that this improved RT-PCR approach could be a fast, simple and specific detection method for IBV.