Add time:09/26/2019         Source:sciencedirect.com

Herein, a novel and convenient electrogenerated chemiluminescence (ECL) biosensing method for the quantitative detection of 5-HYDROXYMETHYLCYTOSINE (cas 1123-95-1) (5-hmC) in genomic DNA was developed by employing thiolated anchor probe DNA to immobilize 5-hydroxymethylcytosine double-stranded DNA (5-hmC-dsDNA) and a ruthenium(II) complex-tagged reporter probe as an ECL probe. Briefly, the 5-hmC-dsDNA was firstly oxidized to 5-formylcytosine dsDNA (5-fC-dsDNA) by KRuO4 in high yields. Subsequently, the thiolated anchor probe DNA is employed to help immobilize the 5-fC-dsDNA onto the gold electrode surface via the Au-S bond. An ECL probe DNA, labeled with a signal transducer − Ru(bpy)2(cbpy)NHS, hybridized with the anchor probe DNA on the electrode, resulting in an intensive ECL emission. The obtained ECL intensity was linearly proportional to the content of 5-hmC in the range from 0.0090% to 0.5761%, with the limit of detection calculated to be 0.0036% (S/N = 3). Finally, we applied the developed 5-hmC detection strategy in various mice tissues. It demonstrates that the content of 5-hmC measured by this method in different mice tissues vary significantly: 5-hmC is highest in brain and lowest in spleen. The success in assaying the mice tissues samples signifies its great promise in a broad range of practical applications for the quantitative detection of 5-hmC in genomic DNA.

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