Examination of the specificity of DNA methylation profiling techniques towards 5-methylcytosine and 5-hydroxymethylcytosine.
Examination of the specificity of DNA methylation profiling techniques towards 5-methylcytosine and 5-hydroxymethylcytosine.
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DOI：
10.1093/nar/gkq223
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发表时间：
2010-06
影响因子：
14.9
通讯作者：
Pfeifer GP
中科院分区：
文献类型：
--
作者：
Jin SG;Kadam S;Pfeifer GP
DNA胞嘧啶-5甲基化是一种被广泛研究的表观遗传途径，与基因表达调控和疾病发病机制有关。已经开发了不同的技术来检测5-甲基胞嘧啶(5mC)在基因组特定序列中的分布。最近，在某些哺乳动物组织中检测到大量的5-羟甲基胞嘧啶(5hmC)，很可能来自TET1对5mC的酶催化氧化。在这里，我们检查了几种常用的DNA甲基化分析方法区分5mC和5hmC的能力。我们表明，基于亚硫酸氢钠处理DNA的技术不能区分这两个修饰的碱基。相反，基于抗5mC抗体的免疫沉淀技术(甲基化DNA免疫沉淀，MeDIP)或基于与甲基化CpG序列结合的蛋白质的技术(例如，甲基化CpG岛恢复试验，MIRA)不能检测到5hmC，并且对5mC是特异的，除非这两个修饰碱基出现在同一DNA片段中。我们还报道了包括MBD1、MBD2和MBD4在内的几种甲基CpG结合蛋白不能与含有5hmC的序列结合。5hmC的选择性测绘将需要开发独特的工具来检测这个修饰的碱基。
DNA cytosine-5 methylation is a well-studied epigenetic pathway implicated in gene expression control and disease pathogenesis. Different technologies have been developed to examine the distribution of 5-methylcytosine (5mC) in specific sequences of the genome. Recently, substantial amounts of 5-hydroxymethylcytosine (5hmC), most likely derived from enzymatic oxidation of 5mC by TET1, have been detected in certain mammalian tissues. Here, we have examined the ability of several commonly used DNA methylation profiling methods to distinguish between 5mC and 5hmC. We show that techniques based on sodium bisulfite treatment of DNA are incapable of distinguishing between the two modified bases. In contrast, techniques based on immunoprecipitation with anti-5mC antibody (methylated DNA immunoprecipitation, MeDIP) or those based on proteins that bind to methylated CpG sequences (e.g. methylated-CpG island recovery assay, MIRA) do not detect 5hmC and are specific for 5mC unless both modified bases occur in the same DNA fragment. We also report that several methyl-CpG binding proteins including MBD1, MBD2 and MBD4 do not bind to sequences containing 5hmC. Selective mapping of 5hmC will require the development of unique tools for the detection of this modified base.
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影响因子：
5
作者：
Rauch, T;Pfeifer, GP
通讯作者：
Pfeifer, GP
DOI：
10.1073/pnas.97.26.14183
发表时间：
2000-12-19
影响因子：
11.1
作者：
Rusmintratip, V;Sowers, LC
通讯作者：
Sowers, LC
DOI：
10.1126/science.1170116
发表时间：
2009-05-15
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Tahiliani M;Koh KP;Shen Y;Pastor WA;Bandukwala H;Brudno Y;Agarwal S;Iyer LM;Liu DR;Aravind L;Rao A
通讯作者：
Rao A
影响因子：
14.8
作者：
Clark, Susan J.;Statham, Aaron;Frommer, Marianne
通讯作者：
Frommer, Marianne
影响因子：
14.9
作者：
CLARK, SJ;HARRISON, J;FROMMER, M
通讯作者：
FROMMER, M