| 荧光定量RT-PCR检测H5亚型禽流感病毒方法的建立与验证 |
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| 引用本文： | 曹军平,陆桂平,徐向明,顾敏,徐全刚,刘武杰,彭大新,刘秀梵.荧光定量RT-PCR检测H5亚型禽流感病毒方法的建立与验证[J].中国畜牧兽医,2011,38(11):169-174. |
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| 作者姓名： | 曹军平 陆桂平 徐向明 顾敏 徐全刚 刘武杰 彭大新 刘秀梵 |
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| 作者单位： | 1. 江苏畜牧兽医职业技术学院,江苏泰州 225300;2. 扬州大学兽医学院,江苏扬州 225009 |
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| 基金项目： | 国家自然科学基金重点项目(30630048); 江苏省农业三项工程项目(SX(2008)092) |
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| 摘 要： | 根据H5亚型禽流感病毒HA基因上的保守序列,设计合成引物,以H5亚型禽流感病毒HA基因重组质粒为标准品绘制标准曲线,建立了荧光定量逆转录聚合酶链反应检测方法。结果表明,本试验建立的标准曲线循环阈值(Ct值)与模板浓度具有良好的线性关系,相关系数为0.995,灵敏度约为8拷贝/μL,相当于8个AIV颗粒,对新城疫病毒和其他禽病病毒无交叉反应,特异性好、重复性佳,为H5亚型禽流感病毒检测提供了一种特异、敏感、快速、低成本、高通量、生物安全性好的定量检测方法。对178份临床泄殖腔棉拭子样品的检测,该方法结果与经典病毒分离方法符合率大于90.0%,在禽流感病毒临床样品快速筛检、流行病学监测等方面显示了良好的应用前景。
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| 关 键 词： | H5亚型禽流感病毒 荧光定量RT-PCR 建立 验证 |
| 收稿时间： | 2011-04-04 |
Development and Validation of Real-time RT-PCR Assay for Detection of Avian Influenza Virus H5 Subtype |
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| CAO Jun-ping,LU Gui-ping,XU Xiang-ming,GU Min,XU Quan-gang,LIU Wu-jie,PENG Da-xin,LIU Xiu-fan.Development and Validation of Real-time RT-PCR Assay for Detection of Avian Influenza Virus H5 Subtype[J].China Animal Husbandry & Veterinary Medicine,2011,38(11):169-174. |
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| Authors: | CAO Jun-ping LU Gui-ping XU Xiang-ming GU Min XU Quan-gang LIU Wu-jie PENG Da-xin LIU Xiu-fan |
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| Institution: | 1. Jiangsu Vocational and Technical College of Animal Husbandry and Veterinary,Taizhou 225300,China;2. College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China |
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| Abstract: | According to the conservative region of H5 AIV HA gene,the primers were devised and synthesized.A serial 10 fold dilutions positive plasmid was prepared and used for standard.The standard curve revealed the linear relationship between Ct(cycle threshold) and template concentration with a good correlation(R2=0.995).The RRT-PCR method for the detection of AIV was highly specific and sensitive,and it could be used for rapid quantitative detection of AIV H5 subtype.No cross-reaction was detected against other a... |
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| Keywords: | AIV H5 subtype real-time RT-PCR development validation |
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