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DNA Modification Detection Methods pp 13–21Cite as

Determination of Cytosine Modifications in DNA by Chemical Labeling-Mass Spectrometry Analysis

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Abstract

In active DNA demethylation, DNA cytosine methylation (5-methylcytosine, 5-mC) can be converted to 5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-fC), and 5-carboxylcytosine (5-caC) by ten-eleven translocation (TET) proteins. These cytosine derivatives play important functions in various biological processes. 5-HmC, 5-fC, and 5-caC in genomic DNA are generally present in low abundance, thus making the quantification of these DNA modifications a challenging task. Here, we developed a method that is capable of determining all the four cytosine modifications in genomic DNA by 2-bromo-1-(4-dimethylamino-phenyl)-ethanone (BDAPE) labeling in combination with liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) analysis. This method enables the sensitive and simultaneous detection of 5-mC, 5-hmC, 5-fC, and 5-caC in genomic DNA.

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Acknowledgements

The work is supported by the National Natural Science Foundation of China (22074110, 21672166, 21721005).

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Authors and Affiliations

  1. Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Sauvage Center for Molecular Sciences, Department of Chemistry, Wuhan University, Wuhan, People’s Republic of China

    Qing-Yun Cheng & Bi-Feng Yuan

Authors
  1. Qing-Yun Cheng
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  2. Bi-Feng Yuan
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Correspondence to Bi-Feng Yuan .

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Editors and Affiliations

  1. Department of Chemistry, Wuhan University, Wuhan, China

    Bi-Feng Yuan

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Cheng, QY., Yuan, BF. (2022). Determination of Cytosine Modifications in DNA by Chemical Labeling-Mass Spectrometry Analysis. In: Yuan, BF. (eds) DNA Modification Detection Methods . Springer Protocols Handbooks. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1229-3_2

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  • DOI: https://doi.org/10.1007/978-1-0716-1229-3_2

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1228-6

  • Online ISBN: 978-1-0716-1229-3

  • eBook Packages: Springer Protocols

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