College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of Education, Wuhan University, Hubei, Wuhan 430072, PR China.
Talanta. 2013 Dec 15;117:445-8. doi: 10.1016/j.talanta.2013.09.026. Epub 2013 Oct 3.
DNA modification, a significant epigenetic event, largely affects genes' binding with the transcription factors and some other DNA binding proteins. Among DNA modifications, methylation, especially cytosine methylation is of great importance and attracts extensive studies as it leads to the silence of tumor-suppressor gene expression. In this work, a novel combined bisulfite Uracil-DNA glycosylase (UDG) assay has been developed on the basis of bisulfite modification to generate uracil from cytosine, subsequent UDG-mediated uracil elimination and ultimate DNA cleavage in alkaline condition. This strategy can be used to selectively detect exact number and loci of 5-methylcytosine residues regardless of sequence context. Moreover, it provides linear quantitative results of DNA methylation level across a wide range.
DNA 修饰是一种重要的表观遗传事件，它主要影响基因与转录因子和其他一些 DNA 结合蛋白的结合。在 DNA 修饰中，甲基化，尤其是胞嘧啶甲基化非常重要，它导致肿瘤抑制基因表达沉默，因此吸引了广泛的研究。在这项工作中，我们基于亚硫酸氢盐修饰开发了一种新的联合亚硫酸盐尿嘧啶-DNA 糖基化酶（UDG）测定法，从胞嘧啶中生成尿嘧啶，随后在碱性条件下由 UDG 介导的尿嘧啶消除和最终的 DNA 断裂。该策略可用于选择性检测 5-甲基胞嘧啶残基的确切数量和位置，而与序列背景无关。此外，它还提供了广泛范围内 DNA 甲基化水平的线性定量结果。
