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DNA Modification Detection Methods pp 115–125Cite as

Determination of N6-Methyladenine in DNA of Mammals and Plants by Dpn I Digestion Combined with Size-Exclusion Ultrafiltration and Mass Spectrometry Analysis

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Abstract

DNA adenine methylation (N6-methyladenine, 6mA) widely exists in bacteria and regulates various biological processes including gene transcription and restriction-modification system. In recent years, it has been reported that 6mA is also present in genomic DNA of higher eukaryotes. However, the accurate determination of 6mA in DNA of higher eukaryotes is challenging owing to the potential contamination of bacterial DNA that normally contains abundant 6mA. Therefore, unambiguous detection and quantification method is required for the accurate analysis of 6mA in DNA of higher eukaryotes. We developed a method by Dpn I digestion combined with size-exclusion ultrafiltration and mass spectrometry analysis to determinate 6mA in DNA of higher eukaryotes, including mammals and plants. The contamination of potential bacterial DNA can be efficiently removed from the extracted eukaryotic DNA, which enables the accurate and confident determination of 6mA in DNA of higher eukaryotes.

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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

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  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 N6-Methyladenine in DNA of Mammals and Plants by Dpn I Digestion Combined with Size-Exclusion Ultrafiltration and 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_11

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  • DOI: https://doi.org/10.1007/978-1-0716-1229-3_11

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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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