猪圆环病毒2型环介导等温扩增(LAMP)检测方法的研究  

Research of loop-mediated isothermal amplification assay for the detection of porcine circovirus type 2

作　　者：何逸民[1] 邹国秋[1] 张得玉[1] 罗玉均[1] 曹宗喜[1] 何冉娅[1] 秦宏阳[1] 孔留五[1] 张桂红[1] 

机构地区：[1]华南农业大学兽医学院,广东广州510642

出　　处：《中国兽医杂志》2009年第12期7-10,共4页Chinese Journal of Veterinary Medicine

基　　金：国家生猪现代农业产业技术体系基金(hycytx009);广东省自然科学基金(5006678);农业部制标项目

摘　　要：本研究介绍了一种便捷、灵敏而又特异的环介导逆转录等温扩增(LAMP)基因检测技术,该技术分别使用特异对应于靶序列中2对特殊引物,并在Bst DNA聚合酶的作用下对靶序列进行等温核酸扩增反应。本研究对LAMP反应体系和反应条件的优化,结果表明PCV-2 LAMP在63℃1 h内能够成功的检测猪圆环病毒2型基因;敏感性和特异性试验表明,本研究能够特异的检测PCV-2并且其敏感度可以达到10个拷贝的DNA分子,初步研究LAMP的阳性检出率与PCR的阳性检出率比较结果为94%符合。以上结果证明,LAMP扩增技术是一种检测程序简便、灵敏度和特异性较高的基因检测手段,在猪圆环病毒2型的快速检测方面具有一定的开发潜力。A simple,sensitive LAMP（loop-mediated isothermal amplification） method for the detection of porcine circovirus type 2（PCV2）DNA is described.The method employs a set of four specially designed primers that recognize distinct sequences of the target for sensitive,specific amplification of nucleicacid under isothermal conditions.In this study,through optimization of reaction condition and reaction system,PCV2 LAMP could successfully detect PCV2 in 63℃ per hour;The test of sensitivity and sensitivity indicated that this method was able to detect PCV2 specially and achieved a sensitivity of 10 copy DNA molecules. Initial investigation showed that the positive detection rate of LAMP was 100% comparing the positive detection rate of PCR. Thus, we could conclude that this established LAMP assay has a potential application tO the rapid detection of PCV2.

关 键 词：猪圆环病毒2型(PCV-2) 环介导等温扩增(LAMP) 敏感性 特异性 快速检测 

分 类 号：S852.659.2[农业科学—基础兽医学]