Zhou Shun, Gao Zhi-Xin, Zhang Min, Liu Dan-Yang, Zhao Xin-Peng, Liu Yong
Marine Science and Engineering College, Qingdao Agricultural University, Qingdao, 266109 People's Republic of China.
College of Fisheries, Qingdao Ocean University, Qingdao, 266100 People's Republic of China.
Springerplus. 2016 Jul 16;5(1):1104. doi: 10.1186/s40064-016-2760-x. eCollection 2016.
A quadruplex loop-mediated isothermal amplification (LAMP) method was developed to detect four Vibrio species, including Vibrio ichthyoenteri, Vibrio parahaemolyticus, Vibrio scophthalmi, and Vibrio vulnificus, simultaneously. Four sets of species-specific primers were designed with different restriction sites contained in the inner primers. The quadruplex LAMP method could distinguish four Vibrio species via the subsequent restriction enzyme analysis. The sensitivity of the quadruplex LAMP method were 10(2)-10(3) times higher than the sensitivity of conventional PCR. V. scophthalmi, V. vulnificus, V. parahaemolyticus and V. ichthyoenteri could be detected in the different tissues of the infected fish by the quadruplex LAMP method simply and conveniently through using SYBR Green I to facilitate visual inspection of the LAMP products. The method we developed in this study could be a simple and convenient diagnostic tool for field detection of Vibrio infection in fish.
开发了一种四重环介导等温扩增（LAMP）方法，用于同时检测四种弧菌，包括鱼肠道弧菌、副溶血性弧菌、大菱鲆弧菌和创伤弧菌。设计了四组物种特异性引物，内引物中包含不同的限制性位点。四重LAMP方法可通过随后的限制性酶切分析区分四种弧菌。四重LAMP方法的灵敏度比传统PCR的灵敏度高10（2）-10（3）倍。通过使用SYBR Green I便于对LAMP产物进行目视检查，四重LAMP方法可以简单方便地在感染鱼的不同组织中检测到大菱鲆弧菌、创伤弧菌、副溶血性弧菌和鱼肠道弧菌。我们在本研究中开发的方法可能是一种简单方便的诊断工具，用于现场检测鱼类中的弧菌感染。
