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基于颜色判定的环介导恒温扩增法快速检测副溶血性弧菌

DOI: 10.1360/052011-534, PP. 1037-1042

Keywords: tlh基因,DNA介导恒温核酸扩增法,快速检测,副溶血性弧菌

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Abstract:

利用DNA环介导恒温核酸扩增法(LAMP)针对副溶血性弧菌特异基因tlh基因设计4条引物,通过引物特异性识别tlh基因上的6个独立区域来快速检测副溶血性弧菌.LAMP反应的过程中会产生白色沉淀焦磷酸镁,故可以通过监测浊度来判定反应结果.实时浊度仪监测反应结果表明,LAMP反应在60~65℃恒温条件下50min内完成;如果在反应前添加羟基萘酚兰(HNB),蓝色的阳性结果很明显区别于紫色阴性结果;LAMP方法的最低检出限为9.74pg/μL,PCR方法最低检出限为97.4pg/μL,LAMP方法检测灵敏度是PCR方法检测灵敏度的10倍,且具有良好的特异性.LAMP方法用于快速检测副溶血性弧菌具有检测过程简单、实验装置简便、反应结果肉眼可辨别、灵敏度高和特异性强的特点,所以LAMP方法检测副溶血性弧菌特别适合用于现场和基层检疫及医疗单位的快速诊断.

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