一种基于HRM技术的快速DNA甲基化检测方法

A novel and reliable method of DNA methylation analysis based on HRM technology

目的:建立经济可靠的基于MS-HRM技术的检测DNA甲基化的方法。方法:采用简单的沉淀法纯化回收Bis-DNA(重亚硫酸钠处理后的DNA),首次采用picogreen为MS-HRM分析染料,并用新建立的方法检测结直肠癌患者组织SEPT9启动子甲基化水平。结果:新方法的回收率在63%以上,纯度、得率都显著优于经典方法中的Promega Wizard DNA Clean-up System,Picogreen染料可检测到低至0.5%甲基化。共检测34例结直肠癌组织,其中SEPT9异常甲基化的组织33例,占97%,检出率高于文献报导。结论:本文建立的方法简单快速、灵敏度高、重复性好、经济、可操作性强的优点。

Objective：To establish a reliable and economic method of DNA methylation analysis based on the High Resolution Melting.Methods：A novel Bis-DNA（bisulfite treated DNA）purification method was established by using ethanol.Picogreen was used as the MS-HRM dye in the analysis.To verify the method,methylation of SEPT9 promotor of colorectal tumor tissue was analyzed.Results：The recovery of DNA purification following bisulfite treatment by the new method was more than 63%,which was much better than Promega Wizard DNA Clean-up System.The sensitivity of HRM using Picogreen dye was as high as 0.5%.Abnormal methylation of SEPT9 promotor was observed in 97% of colorectal tissue samples（33/34）,the detection rate was higher than reported.Conclusion：The new method can be applied to analyze DNA methylation,and it is a reliable and economic method.