Rapid and sensitive detection of Singapore grouper iridovirus by loop-mediated isothermal amplification
Rapid and sensitive detection of Singapore grouper iridovirus by loop-mediated isothermal amplification
复制标题
DOI：
10.1111/j.1365-2672.2008.03761.x
复制
发表时间：
2008-08-01
影响因子：
4
通讯作者：
Qin, Q. W.
中科院分区：
文献类型：
--
作者：
Mao, X. L.;Zhou, S.;Qin, Q. W.
目的：本文的目的是开发一种环介导等温扩增（LAMP）方法，用于快速、灵敏且廉价地检测石斑鱼（GP）、石斑鱼中的新加坡石斑鱼虹彩病毒（SGIV）。方法和结果：通过针对 SGIV ORF-014L 设计了一组六种特异性引物。使用 Bst DNA 聚合酶大片段，在 65°C 的简单水浴中最快 20 分钟即可扩增目标 DNA。检测限约为 0.02 fg（相当于 6.3 个拷贝）的质粒 ORF-014L。 LAMP产品可以用三种不同的方法来判断。与其他七种水生动物病毒没有交叉反应，表明 LAMP 具有高度特异性。 LAMP方法可有效检测病毒感染的GP细胞和GP组织中的SGIV。结论：本研究描述的LAMP是一种廉价、灵敏、特异和快速的检测细胞和GP组织中SGIV的方案。研究意义和影响：所开发的LAMP方法可以简单地应用于现场条件和实验室操作，用于特异性检测SGIV感染。
Aims: The aim of this paper was to develop a loop-mediated isothermal amplification (LAMP) method for rapid, sensitive and inexpensive detection of Singapore grouper iridovirus (SGIV) in grouper (GP), Epinephelus sp.Methods and Results: A set of six specific primers was designed by targeting the SGIV ORF-014L. With Bst DNA polymerase large fragment, the target DNA can be amplified as early as 20 min at 65 degrees C in a simple water bath. The detection limit is about 0.02 fg (equivalent to 6.3 copies) of plasmid ORF-014L. LAMP products could be judged with three different methods. There were no cross-reactions with seven other aquatic animal viruses indicating high specificity of the LAMP. The LAMP method was applied to detect SGIV in virus-infected GP cells and GP tissues effectively.Conclusions: The LAMP described in this study is a cheap, sensitive, specific and rapid protocol for the detection of SGIV in cells and in GP tissues.Significance and Impact of the Study: The developed LAMP method can be simply applied both in field condition and in laboratory operation for specific detection of SGIV infection.