GSE49546
RRHP : a tag-based approach for 5-hydroxymethylcytosine mapping at single-site resolution
We present here a novel approach called Reduced Representation 5-Hydroxymethylcytosine Profiling (RRHP), which exploits ß-glucosyltransferase (ß-GT) to inhibit restriction digestion at adapters ligated to a genomic library, such that only fragments presenting glucosylated 5hmC residues at adapter junctions will be amplified and sequenced. This assay profiles 5hmC sites with single-base resolution in a strand-specific manner. The absence of harsh chemical conversion steps allows for sequencing of native DNA with less inputs, enhancing both sequencing quality and mapping efficiency. Most importantly, the method proves highly reproducible and is a positive display method, sensitive enough to interrogate 5hmC sites with low abundance. When combined with existing RRBS data, it allows simultaneous comparison of 5mC and 5hmC at specific site. Overall design: developing a new assay for genomic profiling of 5hmC