Bisulfite-free direct detection of 5-methylcytosine and 5-hydroxymethylcytosine at base resolution

Nature Biotechnology
Yibin LiuChun-Xiao Song

Abstract

Bisulfite sequencing has been the gold standard for mapping DNA modifications including 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) for decades1-4. However, this harsh chemical treatment degrades the majority of the DNA and generates sequencing libraries with low complexity2,5,6. Here, we present a bisulfite-free and base-level-resolution sequencing method, TET-assisted pyridine borane sequencing (TAPS), for detection of 5mC and 5hmC. TAPS combines ten-eleven translocation (TET) oxidation of 5mC and 5hmC to 5-carboxylcytosine (5caC) with pyridine borane reduction of 5caC to dihydrouracil (DHU). Subsequent PCR converts DHU to thymine, enabling a C-to-T transition of 5mC and 5hmC. TAPS detects modifications directly with high sensitivity and specificity, without affecting unmodified cytosines. This method is nondestructive, preserving DNA fragments over 10 kilobases long. We applied TAPS to the whole-genome mapping of 5mC and 5hmC in mouse embryonic stem cells and show that, compared with bisulfite sequencing, TAPS results in higher mapping rates, more even coverage and lower sequencing costs, thus enabling higher quality, more comprehensive and cheaper methylome analyses.

References

Feb 6, 2007·Bioorganic & Medicinal Chemistry Letters·Kazuo Tanaka, Akimitsu Okamoto
Oct 17, 2007·Annual Review of Medicine·Einav Nili Gal-YamPeter A Jones
May 20, 2009·Bioinformatics·Heng Li, Richard Durbin
Jan 30, 2010·Bioinformatics·Aaron R Quinlan, Ira M Hall
May 11, 2010·Nature Methods·Benjamin A FlusbergStephen W Turner
Jan 12, 2011·Nature Biotechnology·James T RobinsonJill P Mesirov
Apr 16, 2011·Bioinformatics·Felix Krueger, Simon R Andrews
Dec 27, 2011·Bioinformatics·Weichun HuangGabor T Marth
Sep 8, 2012·Nature·UNKNOWN ENCODE Project Consortium
Jun 14, 2013·Journal of the American Chemical Society·Xingyu LuChuan He
Jul 6, 2013·Science·Ryan ListerJoseph R Ecker
Nov 6, 2013·Proceedings of the National Academy of Sciences of the United States of America·K C Allen ChanY M Dennis Lo
May 3, 2014·Cold Spring Harbor Perspectives in Biology·En Li, Yi Zhang
Jul 22, 2014·Nature Methods·Sébastien A SmallwoodGavin Kelsey
Apr 2, 2015·Proceedings of the National Academy of Sciences of the United States of America·June E PaisLana Saleh
Apr 22, 2015·Cancer Genetics·Aparna Vasanthakumar, Lucy A Godley
Feb 13, 2018·Nature Biotechnology·Miten JainMatthew Loose
Apr 19, 2018·PloS One·Amanda RaineJessica Nordlund
Oct 9, 2018·Nature Biotechnology·Emily K SchutskyRahul M Kohli

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Citations

Dec 18, 2019·Essays in Biochemistry·Ksenia SkvortsovaPhillippa Taberlay
Oct 12, 2019·Journal of Human Genetics·Liu Xu, Masahide Seki
Jul 14, 2020·Human Molecular Genetics·Martin SillDaniel B Lipka
Aug 25, 2020·Integrative and Comparative Biology·Arild Husby
Aug 14, 2020·Chembiochem : a European Journal of Chemical Biology·Jenna N BeyerGeorge M Burslem
Feb 6, 2020·Current Environmental Health Reports·Joseph Kochmanski, Alison I Bernstein
Dec 6, 2019·Frontiers in Genetics·Warwick J LockeJason P Ross
Jun 26, 2020·Genome Medicine·Florian ErgerJanine Altmüller
Jun 4, 2020·Epigenetics & Chromatin·Stephany OrjuelaMark D Robinson
Jul 6, 2020·Nucleic Acids Research·Tajda KlobučarMelanie Eckersley-Maslin
Oct 17, 2019·Cells·Marta BorchielliniAnnalisa Di Ruscio
Dec 13, 2019·Dose-response : a Publication of International Hormesis Society·Fan JiangMingming Yang
Oct 10, 2020·Nature Communications·Yinghua HuangBaoming Qin
Nov 22, 2020·Scientific Reports·Fanny WangAdam R Hall
Dec 19, 2020·Biomolecules·Francesca GalardiMatteo Benelli
Apr 8, 2020·Computational and Structural Biotechnology Journal·Li-Qian ChenGuan-Zheng Luo
Sep 11, 2019·Molecular Metabolism·Marie LohJohn Campbell Chambers
Jan 27, 2021·Proceedings of the National Academy of Sciences of the United States of America·O Y Olivia TseY M Dennis Lo
Mar 5, 2021·Epigenomics·Ying TaoXiaofeng Dai
May 1, 2021·Journal of Cardiovascular Development and Disease·Joshua C PetersonMarco C DeRuiter
May 13, 2021·Epigenetics : Official Journal of the DNA Methylation Society·Daniel BeckMichael K Skinner
May 18, 2021·Frontiers in Oncology·Anastasia P KovalDmitry S Shcherbo
May 22, 2021·Nucleic Acids Research·Yoshitaka SakamotoMasahide Seki
Jun 17, 2021·Nature Biomedical Engineering·Imran S Haque
Jul 12, 2021·Seminars in Perinatology·Robert C PeeryLanlan Shen

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Methods Mentioned

BETA
PCR
WGBS
Illumina sequencing
pull-down
electrophoresis

Software Mentioned

caller
Bismark
python3
asTair
MOD
Matlab
R
WGBS
samtools
samtools mpileup

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