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Nat Biotechnol. 2018 Oct 8. doi: 10.1038/nbt.4204. [Epub ahead of print]

Nondestructive, base-resolution sequencing of 5-hydroxymethylcytosine using a DNA deaminase.

Author information

1
Department of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
2
Department of Genetics, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
3
Department of Microbiology, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
4
Penn Epigenetics Institute, University of Pennsylvania, Philadelphia, Pennsylvania, USA.

Abstract

Here we present APOBEC-coupled epigenetic sequencing (ACE-seq), a bisulfite-free method for localizing 5-hydroxymethylcytosine (5hmC) at single-base resolution with low DNA input. The method builds on the observation that AID/APOBEC family DNA deaminase enzymes can potently discriminate between cytosine modification states and exploits the non-destructive nature of enzymatic, rather than chemical, deamination. ACE-seq yielded high-confidence 5hmC profiles with at least 1,000-fold less DNA input than conventional methods. Applying ACE-seq to generate a base-resolution map of 5hmC in tissue-derived cortical excitatory neurons, we found that 5hmC was almost entirely confined to CG dinucleotides. The whole-genome map permitted cytosine, 5-methylcytosine (5mC) and 5hmC to be parsed and revealed genomic features that diverged from global patterns, including enhancers and imprinting control regions with high and low 5hmC/5mC ratios, respectively. Enzymatic deamination overcomes many challenges posed by bisulfite-based methods, thus expanding the scope of epigenome profiling to include scarce samples and opening new lines of inquiry regarding the role of cytosine modifications in genome biology.

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