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Genome-wide mapping of endogenous G-quadruplex DNA structures by chromatin immunoprecipitation and high-throughput sequencing

Nature Protocols volume 13, pages 551–564 (2018)Cite this article

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Abstract

G-rich DNA sequences can form four-stranded G-quadruplex (G4) secondary structures and are linked to fundamental biological processes such as transcription, replication and telomere maintenance. G4s are also implicated in promoting genome instability, cancer and other diseases. Here, we describe a detailed G4 ChIP-seq method that robustly enables the determination of G4 structure formation genome-wide in chromatin. This protocol adapts traditional ChIP-seq for the detection of DNA secondary structures through the use of a G4-structure-specific single-chain antibody with refinements in chromatin immunoprecipitation followed by high-throughput sequencing. This technology does not require expression of the G4 antibody in situ, enabling broad applicability to theoretically all chromatin sources. Beginning with chromatin isolation and antibody preparation, the entire protocol can be completed in <1 week, including basic computational analysis.

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Figure 1: General G4 ChIP-seq workflow.
Figure 3: BG4 quality assessment.
Figure 2: Expected results from a typical G4 ChIP-seq experiment.

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Acknowledgements

We acknowledge support from the University of Cambridge and Cancer Research UK. The Balasubramanian laboratory is supported by core funding from Cancer Research UK (C14303/A17197). S.B. is a Senior Investigator of the Wellcome Trust (grant no. 099232/z/12/z). J.S. is a Marie Curie Fellow of the European Union (747297-QAPs-H2020-MSCA-IF-2016).

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Author notes

  1. Robert Hänsel-Hertsch and Jochen Spiegel: These authors contributed equally to this work.

Authors and Affiliations

  1. Cancer Research UK, Cambridge Institute, Li Ka Shing Centre, Cambridge, UK

    Robert Hänsel-Hertsch, Jochen Spiegel, Giovanni Marsico, David Tannahill & Shankar Balasubramanian

  2. Department of Chemistry, University of Cambridge, Cambridge, UK

    Jochen Spiegel & Shankar Balasubramanian

  3. School of Clinical Medicine, University of Cambridge, Cambridge, UK

    Shankar Balasubramanian

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  1. Robert Hänsel-Hertsch
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Contributions

R.H.-H. developed the G4 ChIP-seq method. R.H.-H. and J.S. performed the experiments. R.H.-H., J.S. and G.M. performed bioinformatics analysis. All authors interpreted the data and wrote the manuscript.

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Correspondence to Shankar Balasubramanian.

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Hänsel-Hertsch, R., Spiegel, J., Marsico, G. et al. Genome-wide mapping of endogenous G-quadruplex DNA structures by chromatin immunoprecipitation and high-throughput sequencing. Nat Protoc 13, 551–564 (2018). https://doi.org/10.1038/nprot.2017.150

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