Oxidation potential-dependent selective detection of epigenetic 5-hydroxymethylcytosine using nanocarbon film

We developed and used the nanocarbon film electrode to successfully detect epigenetic mark 5-hydroxymethylcytosine (5hmC) through direct oxidation after digesting genomic DNA (gDNA) with nuclease P1. The film showed superior key film properties including a very chemically stable structure and an ultraflat surface (R_a = 0.21 nm) that resulted in better electrode properties such as a wider potential window and lower background current than the widely used carbon electrode. Computational experiments revealed that the theoretical oxidation potential (E_ox) value for 5hmC coincided with that obtained in electrochemical experiments using our nanocarbon film electrode. We successfully detected 5hmC by using this film electrode in combination with HPLC. Due to the difference of each oxidation potential, we achieved the oxidation potential-dependent selective detection of 5hmC in gDNA from excess amounts of unmethylated cytosine with a similar polarity to 5hmC.