Xingyu Lu, Chun-Xiao Song, Keith E. Szulwach
Jun 17, 2013
Citations
1
Influential Citations
105
Citations
Quality indicators
Journal
Journal of the American Chemical Society
Abstract
5-Methylcytosine (5mC) in DNA can be oxidized stepwise to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC) by the TET family proteins. Thymine DNA glycosylase can further remove 5fC and 5caC, connecting 5mC oxidation with active DNA demethylation. Here, we present a chemical modification-assisted bisulfite sequencing (CAB-Seq) that can detect 5caC with single-base resolution in DNA. We optimized 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC)-catalyzed amide bond formation between the carboxyl group of 5caC and a primary amine group. We found that the modified 5caC can survive the bisulfite treatment without deamination. Therefore, this chemical labeling coupled with bisulfite treatment provides a base-resolution detection and sequencing method for 5caC.