Genomic DNA methylation, occurring at the C5 position of cytosine (C) in the 5'-CG-3'sequence (CpG), plays an important role in the regulation of gene transcription.1-4 5-hydroxymethylcytosine (5-hmC) is a naturally occurring nucleobase in mammals that is newly discovered in 2009.5 5-hmC is produced from the oxidation of 5-methylcytosine (5-mC) by the ten-eleven translocation (TET) proteins. Thus, a discriminative determination of 5-hmC from 5-mC is important for epigenetic study.  A novel study to oxidize and selective label (Ox-Label) 5-hydroxymethylcytosine (5-hmC) in native DNA has been conducted, which conveniently distinguishes 5-hmC from 5-methylcytosine (5-mC) using simple and effective processes. A two-step strategy is developed to directly label 5-hmC in DNA using fluorescent dyes. This Ox-Label approach succeeded in determining the distribution of 5-hmC and 5-mC in DNAs. The newly developed method does not require long reaction times, enzymatic digestion or genetic sequencing. Using PAGE and fluorescent measurement, qualitative and quantitative research on 5-hmC in DNA could be achieved conveniently. The use of this method to understand the role of 5-hmC modification at the molecular level requires further investigation. The next challenge will be to increase the sensitivity and accuracy of this method. Thus, our method represents a very promising tool in future DNA methylation study.

Discrimination between 5-hydroxymethylcytosine and 5-methylcytosine in DNA by selective chemical labeling

246th National Meeting (2013)

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