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Chembiochem. 2016 Mar 2;17(5):403-6. doi: 10.1002/cbic.201500646. Epub 2016 Feb 5.

Identification of Sequence Specificity of 5-Methylcytosine Oxidation by Tet1 Protein with High-Throughput Sequencing.

Author information

1
Department of Chemistry, Graduate School of Science, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto-shi, Kyoto, 606-8502, Japan.
2
Department of Chemistry, Graduate School of Science, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto-shi, Kyoto, 606-8502, Japan. hs@kuchem.kyoto-u.ac.jp.
3
Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Yoshida-ushinomiyacho, Sakyo-ku, Kyoto-shi, Kyoto, 606-8501, Japan. hs@kuchem.kyoto-u.ac.jp.

Abstract

Tet (ten-eleven translocation) family proteins have the ability to oxidize 5-methylcytosine (mC) to 5-hydroxymethylcytosine (hmC), 5-formylcytosine (fC), and 5-carboxycytosine (caC). However, the oxidation reaction of Tet is not understood completely. Evaluation of genomic-level epigenetic changes by Tet protein requires unbiased identification of the highly selective oxidation sites. In this study, we used high-throughput sequencing to investigate the sequence specificity of mC oxidation by Tet1. A 6.6×10(4) -member mC-containing random DNA-sequence library was constructed. The library was subjected to Tet-reactive pulldown followed by high-throughput sequencing. Analysis of the obtained sequence data identified the Tet1-reactive sequences. We identified mCpG as a highly reactive sequence of Tet1 protein.

KEYWORDS:

5-hydroxymethylcytosine; 5-methylcytosine; DNA recognition; Tet; high-throughput sequencing; sequence determination

PMID:
26715454
DOI:
10.1002/cbic.201500646
[Indexed for MEDLINE]

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