Base-Resolution Analysis of 5-Hydroxymethylcytosine in the Mammalian Genome

• Authors: Miao Yu, The University of Chicago; Gary C. Hon, University of California; Keith E. Szulwach, Emory University; Chun-Xiao Song, The University of Chicago; Liang Zhang, The University of Chicago; Audrey Kim, University of California; Xuekun Li, Emory University; Qing Dai, The University of Chicago; Beomseok Park, The University of Illinois; Jung-Hyun Min, The University of Illinois; Peng Jin, Emory University; Bing Ren, University of California; Chuan He, The University of Chicago
• Journal Title: Cell
• Volume: Volume 149, Number 6
• Publisher: Elsevier (Cell Press) | 2012-06-08, Pages 1368-1380
• Type of Work: Article | Post-print: After Peer Review
• Publisher DOI: http://doi.org/10.1016/j.cell.2012.04.027
• Subjects: Chemistry, General; Biology, Genetics; Health Sciences, Oncology

Abstract

SUMMARY 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), for mapping 5hmC at base resolution and quantifying the relative abundance of 5hmC as well as 5mC when combined with traditional bisulfite sequencing. 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.