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Efficient Targeted DNA Methylation with dCas9-Coupled DNMT3A-DNMT3L Methyltransferase

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Epigenomics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2577))

Abstract

Epigenome editing is a powerful approach for the establishment of a chromatin environment with desired properties at a selected genomic locus, which is used to influence the transcription of target genes and to study properties and functions of gene regulatory elements. Targeted DNA methylation is one of the most often used types of epigenome editing, which typically aims for gene silencing by methylation of gene promoters. Here, we describe the design principles of EpiEditors for targeted DNA methylation and provide step-by-step guidelines for the realization of this approach. We focus on the dCas9 protein as the state-of-the-art DNA targeting module fused to 10×SunTag as the most frequently used system for editing enhancement. Further, we discuss different flavors of DNA methyltransferase modules used for this purpose including the most specific variants developed recently. Finally, we explain the principles of gRNA selection, outline the setup of the cell culture experiments, and briefly introduce the available options for the downstream DNA methylation data analysis.

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Acknowledgments

Work in the authors’ laboratory has been supported by the BW Foundation (AllEpi, ID09).

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Authors and Affiliations

  1. Institute of Biochemistry and Technical Biochemistry, Department of Biochemistry, University of Stuttgart, Stuttgart, Germany

    Pavel Bashtrykov, Nivethika Rajaram & Albert Jeltsch

Authors
  1. Pavel Bashtrykov
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  2. Nivethika Rajaram
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  3. Albert Jeltsch
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Corresponding authors

Correspondence to Pavel Bashtrykov or Albert Jeltsch .

Editor information

Editors and Affiliations

  1. Institute for Molecular and Cellular Regulation, Gunma University, Maebashi, Gunma, Japan

    Izuho Hatada

  2. Institute for Molecular and Cellular Regulation, Gunma University, Maebashi, Gunma, Japan

    Takuro Horii

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© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

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Bashtrykov, P., Rajaram, N., Jeltsch, A. (2023). Efficient Targeted DNA Methylation with dCas9-Coupled DNMT3A-DNMT3L Methyltransferase. In: Hatada, I., Horii, T. (eds) Epigenomics. Methods in Molecular Biology, vol 2577. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2724-2_12

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  • DOI: https://doi.org/10.1007/978-1-0716-2724-2_12

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2723-5

  • Online ISBN: 978-1-0716-2724-2

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