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Genome-scale DNA methylation mapping of clinical samples at single-nucleotide resolution

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Abstract

Bisulfite sequencing measures absolute levels of DNA methylation at single-nucleotide resolution, providing a robust platform for molecular diagnostics. We optimized bisulfite sequencing for genome-scale analysis of clinical samples: here we outline how restriction digestion targets bisulfite sequencing to hotspots of epigenetic regulation and describe a statistical method for assessing significance of altered DNA methylation patterns. Thirty nanograms of DNA was sufficient for genome-scale analysis and our protocol worked well on formalin-fixed, paraffin-embedded samples.

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Figure 1: Optimizing bisulfite sequencing for genome-scale profiling of human disease samples.

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Acknowledgements

We thank K. Halachev (Max Planck Institute for Informatics) for providing genome annotation files and H. Cedar (The Hebrew University of Jerusalem) for providing the human blood DNA samples. C.B. is supported by a Feodor Lynen Fellowship from the Alexander von Humboldt Foundation. A.M. is supported by the Massachusetts Life Science Center and the Pew Charitable Trusts. The described work was in part funded by US National Institutes of Health grants R01HG004401, U54HG03067 and U01ES017155.

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Author notes

  1. Hongcang Gu and Christoph Bock: These authors contributed equally to this work.

Authors and Affiliations

  1. Broad Institute, Cambridge, Massachusetts, USA

    Hongcang Gu, Christoph Bock, Tarjei S Mikkelsen, Natalie Jäger, Zachary D Smith, Eleni Tomazou, Andreas Gnirke, Eric S Lander & Alexander Meissner

  2. Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts, USA

    Christoph Bock, Natalie Jäger, Zachary D Smith, Eleni Tomazou & Alexander Meissner

  3. Harvard Stem Cell Institute, Cambridge, Massachusetts, USA

    Christoph Bock, Natalie Jäger, Zachary D Smith, Eleni Tomazou & Alexander Meissner

  4. Max Planck Institute for Informatics, Saarbrücken, Germany

    Christoph Bock

  5. Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA

    Eric S Lander

  6. Department of Systems Biology, Harvard Medical School, Boston, Massachusetts, USA

    Eric S Lander

Authors
  1. Hongcang Gu
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  2. Christoph Bock
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  3. Tarjei S Mikkelsen
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  4. Natalie Jäger
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  5. Zachary D Smith
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  6. Eleni Tomazou
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  7. Andreas Gnirke
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  8. Eric S Lander
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  9. Alexander Meissner
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Contributions

H.G., C.B., A.G., E.S.L. and A.M. conceived and designed the experiments; H.G. and E.T. performed the experiments; C.B. analyzed data; T.S.M., N.J. and Z.D.S. contributed materials or analysis tools; and H.G., C.B. and A.M. wrote the paper.

Corresponding author

Correspondence to Alexander Meissner.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–2 and Supplementary Table 1 (PDF 822 kb)

Supplementary Note

Source code of Epigenome pipeline package with documentation and demonstration data. (ZIP 88748 kb)

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Gu, H., Bock, C., Mikkelsen, T. et al. Genome-scale DNA methylation mapping of clinical samples at single-nucleotide resolution. Nat Methods 7, 133–136 (2010). https://doi.org/10.1038/nmeth.1414

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