Abstract
Studies of epigenetic modifications would benefit from improved methods for high-throughput methylation profiling. We introduce two complementary approaches that use next-generation sequencing technology to detect cytosine methylation. In the first method, we designed ∼10,000 bisulfite padlock probes to profile ∼7,000 CpG locations distributed over the ENCODE pilot project regions and applied them to human B-lymphocytes, fibroblasts and induced pluripotent stem cells. This unbiased choice of targets takes advantage of existing expression and chromatin immunoprecipitation data and enabled us to observe a pattern of low promoter methylation and high gene-body methylation in highly expressed genes. The second method, methyl-sensitive cut counting, generated nontargeted genome-scale data for ∼1.4 million HpaII sites in the DNA of B-lymphocytes and confirmed that gene-body methylation in highly expressed genes is a consistent phenomenon throughout the human genome. Our observations highlight the usefulness of techniques that are not inherently or intentionally biased towards particular subsets like CpG islands or promoter regions.
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08 May 2009
In the version of this article initially published, the second affilliation for Yuan Gao was omitted: Department of Computer Science, Virginia Commonwealth University, Richmond, Virginia, USA. The affiliation has been added to the HTML and PDF versions of the article.
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Acknowledgements
We thank Kun Zhang for discussion throughout this work; Wei Lin for help with computational design; Andrew Chess and Ravid Straussman for discussion and critical reading of the manuscript; Harvard Biopolymers Facility for Solexa sequencing; and Harvard Partners Center for Genetics and Genomics for gene expression profiling. This work was supported by the NHGRI-Centers of Excellence in Genomic Science (to G.M.C.).
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M.P.B., J.B.L. and G.M.C. conceived the study, designed the research and wrote the manuscript. M.P.B. and J.B.L. performed experiments and data analysis. Y.G. and B.X. carried out initial Solexa sequencing. J.-H.L. helped with culturing cell lines and isolating DNA/RNA. E.M.L. synthesized the padlock oligos. I.-H.P. and G.Q.D. generated the iPS cell lines.
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E.M.L. is an employee of Agilent Technologies. G.M.C. is involved in eight next-generation sequencing companies. M.P.B., J.B.L. and G.M.C. are named as inventors on a patent application on technologies described in this article.
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Supplementary Figures 1–10; Supplementary Tables 2–5, 7,8 (PDF 6072 kb)
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BSPP data (TXT 2113 kb)
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MSCC data (TXT 60051 kb)
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Ball, M., Li, J., Gao, Y. et al. Targeted and genome-scale strategies reveal gene-body methylation signatures in human cells. Nat Biotechnol 27, 361–368 (2009). https://doi.org/10.1038/nbt.1533
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DOI: https://doi.org/10.1038/nbt.1533
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