Base-resolution analysis of 5-hydroxymethylcytosine in the mammalian genome

Miao Yu, Gary C. Hon, Keith E. Szulwach, Chun Xiao Song, Liang Zhang, Audrey Kim, Xuekun Li, Qing Dai, Yin Shen, Beomseok Park, Jung Hyun Min, Peng Jin, Bing Ren, Chuan He

Research output: Contribution to journalArticlepeer-review

824 Scopus citations


The study of 5-hydroxylmethylcytosines (5hmC) has been hampered by the lack of a method to map it at single-base resolution on a genome-wide scale. Affinity purification-based methods cannot precisely locate 5hmC nor accurately determine its relative abundance at each modified site. We here present a genome-wide approach, Tet-assisted bisulfite sequencing (TAB-Seq), that when combined with traditional bisulfite sequencing can be used for mapping 5hmC at base resolution and quantifying the relative abundance of 5hmC as well as 5mC. Application of this method to embryonic stem cells not only confirms widespread distribution of 5hmC in the mammalian genome but also reveals sequence bias and strand asymmetry at 5hmC sites. We observe high levels of 5hmC and reciprocally low levels of 5mC near but not on transcription factor-binding sites. Additionally, the relative abundance of 5hmC varies significantly among distinct functional sequence elements, suggesting different mechanisms for 5hmC deposition and maintenance.

Original languageEnglish (US)
Pages (from-to)1368-1380
Number of pages13
Issue number6
StatePublished - Jun 8 2012

ASJC Scopus subject areas

  • General Biochemistry, Genetics and Molecular Biology


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