| 羟基萘酚蓝在伪狂犬病病毒环介导等温扩增检测方法中的应用 |
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| 引用本文： | 张侃,蒋菲,张跃伟,李佳禾,郭盼盼,黄书林,吴文学.羟基萘酚蓝在伪狂犬病病毒环介导等温扩增检测方法中的应用[J].农业生物技术学报,2011,19(6). |
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| 作者姓名： | 张侃 蒋菲 张跃伟 李佳禾 郭盼盼 黄书林 吴文学 |
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| 作者单位： | 中国农业大学动物医学院,北京,100193 |
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| 基金项目： | 农业部动物疫情监测与防治项目资助 |
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| 摘 要： | 伪狂犬病毒(Pseudorabies virus,PRV)传统诊断方法具有程序繁琐、周期长、成本高、特异性差等缺点,本实验建立了敏感、特异的PRV的环介导等温扩增(loop-mediated isothermal amplification,LAMP)检测方法,并在此基础上应用金属离子指示剂羟基萘酚蓝(hydroxy naphthol blue,HNB)进行反应结果判定.实验选取保守性较高的gB基因(glycoprotein B,糖蛋白gB)筛选引物,确定反应体系后,可在45 min内完成检测.LAMP方法特异性与PCR方法一致,敏感性是PCR方法的100倍,最低能够检测10个拷贝的目的基因.在HNB的应用试验中,发现LAMP反应体系中,HNB浓度为150 μmol/L时,阴阳性的颜色对比明显且反应的敏感性最高,可用于LAMP扩增产物的判断.临床样本的检测结果表明,PCR方法和LAMP方法的检出率一致.因此,本研究中所建立的LAMP方法是一种高度特异敏感的,可替代PCR方法的检测技术.
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| 关 键 词： | 伪狂犬病病毒(PRV) 环介导等温扩增(LAMP) PCR 羟基萘酚蓝(HNB) |
The Application of Hydroxy Naphthol Blue in Loop-mediated Isothermal Amplification of Pseudorabies virus |
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| Zhang Kan
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Jiang Fei
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Zhang Yuewei
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Li Jiahe
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Guo Panpan
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Huang Shulin
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Wu Wenxue.The Application of Hydroxy Naphthol Blue in Loop-mediated Isothermal Amplification of Pseudorabies virus[J].Journal of Agricultural Biotechnology,2011,19(6). |
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| Authors: | Zhang Kan Jiang Fei Zhang Yuewei Li Jiahe Guo Panpan Huang Shulin Wu Wenxue |
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| Institution: | Zhang Kan Jiang Fei Zhang Yuewei Li Jiahe Guo Panpan Huang Shulin Wu Wenxue* College of Veterinary Medicine,China Agriculture University,Beijing 100193,China |
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| Abstract: | The traditional diagnostic method for Pseudorabies virus(PRV) has the defect of cumbersome procedures,long period,high cost and poor specificity.Thus we built a sensitive and specific loop-mediated isothermal amplification(LAMP) method for the detection of PRV,and used hydroxy naphthol blue(HNB) as indicator to judge the reaction result.The highly conserved gB(glycoprotein B)gene was selected to design primers.The detection could be finished within 45 min after the reaction system was established.Both LAMP ... |
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| Keywords: | Pseudorabies virus(PRV) Loop-mediated isothermal amplification(LAMP) PCR Hydroxy naphthol blue(HNB) |
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