南芥菜花叶病毒RT-PCR扩增产物胶体金层析和酶联检测方法的建立

• 作 者: 曹成 魏梅生 张永江 李桂芬 吴兴泉
• 机 构: 河南工业大学生物工程学院,中国检验检疫科学研究院动植物检疫研究所,
• 出 处: 植物检疫
• 发表日期: 2011年05期
• 关键词: 胶体金层析 南芥菜花叶病毒 胶体金 试纸条 核酸检测 扩增产物 进境植物 双标记 检测 检疫性有害生物

南芥菜花叶病毒是我国进境植物检疫性有害生物,为建立快速灵敏的检疫检测方法,本研究用生物素标记一条引物,用荧光素(或地高辛)标记另一条引物,经RT-PCR扩增产生双标记的扩增产物,建立了PCR扩增产物的胶体金层析检测和ELISA检测方法。胶体金层析检测方法在15min即可获得检测结果,ELISA方法需20h,后者检测的灵敏度可比前者分别提高100倍和5000倍。所建立的2种方法都免去了普通PCR检测的溴化乙锭染色和电泳的过程。

In order to establish a fast and sensitive quarantine detection method, Arabidopsis mosaic virus is a phytosanitary pest inhabiting in China. In this study, one primer was labeled with biotin, another primer was labeled with fluorescein (or digoxin) PCR amplification to generate double-labeled amplification products, established PCR amplification of colloidal gold and ELISA detection methods. Colloidal gold detection method can be obtained in 15min test results, ELISA method requires 20h, the latter detection sensitivity than the former increased 100-fold and 5000-fold, respectively. All the two methods were developed to eliminate the ethidium bromide staining and electrophoresis of common PCR.