Figure 1.

Selective labeling and enrichment of 5hmU in genomic DNA using JGT. (A) The hydroxyl group of 5hmU in duplex DNA can be glycosylated by JGT to form β-d-glucosylhydroxymethyluridine (glucosyl-5hmU), also called base J. (B) Overview of the selective labeling and enrichment strategy for 5hmU. Scheme 1: JGT is utilized to selectively label 5hmU in genomic DNA with N₃-glucose. After addition of the biotin tag through click chemistry, the 5hmU-containing DNA fragments can be enriched by streptavidin-coupled beads allowing detection and sequencing. Scheme 2: JGT is utilized to selectively label 5hmU with glucose allowing subsequent affinity purification using the base JBP1 or antisera against base J. The glucosylated base can also be directly sequenced using SMRT-sequencing technology.