We present a chemical method to selectively tag and enrich thymine modifications, 5-formyluracil (5-fU) and 5-hydroxymethyluracil (5-hmU), found naturally in DNA. Inherent reactivity differences have enabled us to tag 5-fU chemoselectively over its C modification counterpart, 5-formylcytosine (5-fC). We rationalized the enhanced reactivity of 5-fU compared to 5-fC via ab initio quantum mechanical calculations. We exploited this chemical tagging reaction to provide proof of concept for the enrichment of 5-fU containing DNA from a pool that contains 5-fC or no modification. We further demonstrate that 5-hmU can be chemically oxidized to 5-fU, providing a strategy for the enrichment of 5-hmU. These methods will enable the mapping of 5-fU and 5-hmU in genomic DNA, to provide insights into their functional role and dynamics in biology.R.E.H. is supported by The University of Cambridge, F.K. is
supported by the Wellcome Trust, and A.B.S. is supported by
the Herchel Smith Fund. The Balasubramanian group is core-
funded by a Wellcome Trust Senior Investigator Award and by
Cancer Research UK. Departmental NMR facilities are
supported by EPSRC grant EP/K039520/1.This is the final version. It was first published by ACS at http://pubs.acs.org/doi/abs/10.1021/jacs.5b03730
