【摘要】 来源于花椰菜花叶病毒(Cauliflower mosaic virus)的CaMV35S启动子是目前商业化转基因作物中最常用的启动子,对其甲基化水平的测定是研究转基因沉默的重要内容之一。本研究通过甲基化敏感内切酶HpyCH4Ⅳ酶切和实时荧光定量PCR扩增技术,建立了CaMV35S启动子甲基化水平的快速测定方法。利用建立的方法分析了我国安徽省2017年市场的棉花(Gossypium hirsutum)种子,发现部分种子中CaMV35S启动子甲基化处于较高水平。本研究为实现转基因材料中CaMV35S启动子甲基化的快速高通量筛查提供了检测方法。基于相同的原理也可以建立其他转基因元件和DNA序列甲基化水平的快速检测方法。更多还原

【Abstract】 The CaMV35S promoter derived from Cauliflower mosaic virus is the most commonly promoter used in commercialized transgenic crops till now. The analysis of the methylation level of CaMV35S promoter is one of the great aspects for studying the silence of the transgene. In this study, a rapid analysis method for the methylation level of CaMV35S promoter using the digestion of the methylation sensitive endonuclease HpyCH4 Ⅳ and the real-time fluorescence quantitative PCR was established. The methylation level of CaMV35S promoter of the cotton(Gossypium hirsutum) seed samples collected from Anhui province in 2017 was analyzed using this method, and high level of methylation of CaMV35S was found in some of those samples. This study provided a rapid and high throughput detection method for methylation of CaMV35S promoter in transgenic samples. Based on the same principle, rapid detection methods for methylation of the other transgenic elements or DNA sequences in transgenic crops can also be established.更多还原