Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
8340750 | Methods | 2015 | 5 Pages |
Abstract
How DNA demethylation is achieved in mammals is still under extensive investigation. One proposed mechanism is deamination of 5-hydroxymethylcytosine to form 5-hydroxymethyluracil (5hmU), followed by base excision repair to replace the mismatched 5hmU with cytosine. In this process, 5hmU:G mispair serves as a key intermediate and its localization and distribution in mammalian genome could be important information to investigate the proposed pathway. Here we describe a selective labeling method to map mismatched 5hmU. After converting other cytosine modifications to 5-carboxylcytosines, a biotin tag is installed onto mismatched 5hmU through β-glucosyltransferase-catalyzed glucosylation and click chemistry. The enriched 5hmU-containing DNA fragments can be subject to subsequent sequencing to reveal the distribution of 5hmU:G mispair with base-resolution information acquired.
Keywords
5fC5hmUTETBERTET proteins5hmCUDP-GlcTDG5caCDeaminationAPOBEC5mC5-Hydroxymethyluracil5-formylcytosine5-Methylcytosine5-hydroxymethylcytosine5-carboxylcytosineDemethylationThymine DNA glycosylaseβ-Glucosyltransferasebase excision repairten-eleven translocationActivation-induced deaminaseMeSAIDuridine diphosphoglucose
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Authors
Miao Yu, Chun-Xiao Song, Chuan He,