Gene specific-loci quantitative and single-base resolution analysis of 5-formylcytosine by compound-mediated polymerase chain reaction

Chemical Science
Yafen WangXiang Zhou

Abstract

5-Formylcytosine (5fC) is known as one of the key players in the process of active DNA demethylation and displays essential epigenetic functions in mammals. In spite of the blooming development of whole genome sequencing methods for this modified cytosine base, the easily operated gene specific-loci detection of 5fC has rarely been reported. Herein, we present a compound-mediated analysis of the content and site of 5fC by the polymerase chain reaction (PCR) assay. The molecule, namely azi-BP, which can selectively label 5fC and form a huge group through a click chemistry reaction, hindering the amplification activity of Taq DNA polymerase, acts as a "roadblock" and enables the quantitative analysis of 5fC by quantitative polymerase chain reaction (qPCR). The existence of 5fC in several fragment-specific genomic DNAs of mouse embryonic stem cells (mESCs) was successfully demonstrated using this method. In addition, the gene fragment containing 5fC can be easily biotinylated and enriched after labeling with azi-BP. Moreover, after azi-BP incorporation, the loss of the exocyclic 4-amino group of 5fC leads to C-to-T conversion and subsequent pairing with adenine (A) in the PCR, which can accurately identify 5fC sites at single-base...Continue Reading

References

Apr 15, 2011·Nature·Kristine WilliamsKristian Helin
Nov 22, 2011·Nature Methods·Chun-Xiao SongJonas Korlach
Mar 21, 2013·Chemistry : a European Journal·Jianlin HuXiang Zhou
Oct 25, 2013·Nature·Rahul M Kohli, Yi Zhang
Apr 24, 2014·Nature Chemistry·Michael J BoothShankar Balasubramanian
Aug 6, 2014·Chemical Reviews·Michael J BoothShankar Balasubramanian
Dec 17, 2014·Nature Structural & Molecular Biology·Eun-Ang RaiberShankar Balasubramanian
May 7, 2015·Journal of the American Chemical Society·Robyn E HardistyShankar Balasubramanian
Jul 22, 2015·Epigenetics & Chromatin·Achim Breiling, Frank Lyko
Dec 19, 2015·Angewandte Chemie·Biswajit SamantaClaudia Höbartner
Jun 18, 2016·Current Opinion in Chemical Biology·Monica Yun LiuRahul M Kohli
Mar 30, 2017·Chemical Society Reviews·Yuqi ChenXiang Zhou
May 31, 2017·Nature Reviews. Genetics·Xiaoji Wu, Yi Zhang
Jul 1, 2017·Chemical Science·Chaoxing LiuXiang Zhou
Jul 26, 2017·Journal of the American Chemical Society·Fengchao LiChuanzheng Zhou
Sep 13, 2017·Angewandte Chemie·Shaofei JiNatalia Y Tretyakova

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Citations

Feb 28, 2019·Chembiochem : a European Journal of Chemical Biology·Hu ZengChengqi Yi
Oct 22, 2020·Chemical Communications : Chem Comm·Yichuan ZhangAnnamaria Lilienkampf
Jul 16, 2019·DNA Repair·Yingqian Zhang, Chuanzheng Zhou
Jan 21, 2021·Clinical Epigenetics·Anna RugowskaPatryk Konieczny
Jan 23, 2019·Accounts of Chemical Research·Yafen WangXiang Zhou
Nov 18, 2018·Analytical Chemistry·Chaoxing LiuXiang Zhou
Jul 27, 2018·Analytical Chemistry·Yushu YuanXiang Zhou
Aug 31, 2021·RSC Chemical Biology·Yi DaiYu-Qi Feng
Dec 9, 2021·Chemical Communications : Chem Comm·Jizhou LiuXiang Zhou

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Methods Mentioned

BETA
PCR
electrophoresis
pull down

Software Mentioned

CLEVER

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