Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine.
Oxidative bisulfite sequencing of 5-methylcytosine and 5-hydroxymethylcytosine.
复制标题
DOI：
10.1038/nprot.2013.115
复制
发表时间：
2013-10
期刊：
影响因子：
14.8
通讯作者：
中科院分区：
文献类型：
--
作者：
为了揭示哺乳动物胞嘧啶修饰5-甲基胞嘧啶（5 mC）和5-羟甲基胞嘧啶（5 hmC）之间的功能和相互作用，需要新的技术和当前技术的进展。为此，我们开发了氧化亚硫酸氢盐测序（oxBs-seq），它可以在基因组DNA中以单碱基分辨率定量定位5 mC和5 hmC标记。在亚硫酸氢盐测序（BS-seq）中，5 mC和5 hmC都被读取为胞嘧啶，因此无法区分;然而，在oxBS-seq中，5 hmC特异性氧化为5-甲酰基胞嘧啶（5 fC）和新形成的5 fC转化为尿嘧啶（在亚硫酸氢盐条件下）意味着5 hmC可以与5 mC区分开来。从单次oxBS-seq运行中获得实际5 mC的阳性读数，并通过与BS-seq运行比较来推断5 hmC水平。在这里，我们描述了一个优化的第二代协议，可以在二维完成。
To uncover the function of and interplay between the mammalian cytosine modifications 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), new techniques and advances in current technology are needed. To this end, we have developed oxidative bisulfite sequencing (oxBs-seq), which can quantitatively locate 5mC and 5hmC marks at single-base resolution in genomic DNA. In bisulfite sequencing (BS-seq), both 5mC and 5hmC are read as cytosines and thus cannot be discriminated; however, in oxBS-seq, specific oxidation of 5hmC to 5-formylcytosine (5fC) and conversion of the newly formed 5fC to uracil (under bisulfite conditions) means that 5hmC can be discriminated from 5mC. a positive readout of actual 5mC is gained from a single oxBS-seq run, and 5hmC levels are inferred by comparison with a BS-seq run. Here we describe an optimized second-generation protocol that can be completed in 2 d.
登录
查看更多内容
影响因子：
14.8
作者：
Zhang, Liang;Lu, Xingyu;Lu, Junyan;Liang, Haihua;Dai, Qing;Xu, Guo-Liang;Luo, Cheng;Jiang, Hualiang;He, Chuan
通讯作者：
He, Chuan
影响因子：
48
作者：
Flusberg, Benjamin A.;Webster, Dale R.;Lee, Jessica H.;Travers, Kevin J.;Olivares, Eric C.;Clark, Tyson A.;Korlach, Jonas;Turner, Stephen W.
通讯作者：
Turner, Stephen W.
影响因子：
3.7
作者：
Globisch D;Münzel M;Müller M;Michalakis S;Wagner M;Koch S;Brückl T;Biel M;Carell T
通讯作者：
Carell T
影响因子：
8.8
作者：
Sun Z;Terragni J;Borgaro JG;Liu Y;Yu L;Guan S;Wang H;Sun D;Cheng X;Zhu Z;Pradhan S;Zheng Y
通讯作者：
Zheng Y
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