Library-free methylation sequencing with bisulfite padlock probes

Abstract

Targeted quantification of DNA methylation allows for interrogation of the most informative loci across many samples quickly and cost-effectively. Here we report improved bisulfite padlock probes (BSPPs) with a design algorithm to generate efficient padlock probes, a library-free protocol that dramatically reduces sample-preparation cost and time and is compatible with automation, and an efficient bioinformatics pipeline to accurately obtain both methylation levels and genotypes from sequencing of bisulfite-converted DNA.

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Figure 1: Library-free BSPP protocol.

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Acknowledgements

We thank A. Feinberg and J. Stamatoyannopoulos for providing informative genomic targets, and E. LeProust (Agilent Technology) for long-oligonucleotide synthesis. This work was funded by grants from US National Institutes of Health (R01 DA025779 and R01 GM097253) to K.Z.; A.G. is sponsored by a California Institute of Regenerative Medicine predoctoral fellowship. N.P. is supported by the Royal Thai Government Scholarship.

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Contributions

K.Z. oversaw the project. D.D. and N.P. performed experiments and bioinformatic analyses. A.G. developed a probe design algorithm. R.S. and A.G. designed probes. H.-L.F. performed sequencing. D.D., N.P., A.G. and K.Z. wrote the manuscript.

Corresponding author

Correspondence to Kun Zhang.

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Competing interests

K.Z. is a paid consultant for Pathogenica Inc. R.S. is a current employee of Illumina Inc.

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Supplementary Figures 1–11 and Supplementary Tables 1–4 (PDF 1364 kb)

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Diep, D., Plongthongkum, N., Gore, A. et al. Library-free methylation sequencing with bisulfite padlock probes. Nat Methods 9, 270–272 (2012). https://doi.org/10.1038/nmeth.1871

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