DNA methylation detection and inhibitor screening based on the discrimination of the aggregation of long and short DNA on a negatively charged indium tin oxide microelectrode.

On the basis of the different diffusivity and existence of electrostatic repulsion between long and short DNA on the negatively changed indium tin oxide (ITO) microelectrode, a simple but sensitive immobilization free solution-phase electrochemical method for DNA methylation detection and inhibitor screening has been developed. Electroactive substance (methylene blue) tagged at the penultimate base T close to the 3'-terminal first, in the absence of DNA methylation, methylene blue-labeled electroactive fragments cannot be generated by Dpn I and results in a weak electrochemical response being detected on the ITO electrode. On the contrary, a remarkable electrochemical response can be achieved by the cleavage in the presence of DNA methylation since methylene blue-labeled electroactive fragments can be generated and aggregate on the ITO electrode. The proposed system does not need complex operation procedures such as bisulfite treatment, PCR amplification, and electrode immobilization. Six ITO microelectrodes had been assembled on the same microchip, which can achieve the parallel detection of the same sample and improve the experimental efficiency of drug screening. The system was used to conveniently and specifically monitor the change of the DNA methylation level with high sensitivity and selectivity. The proposed system has the potential application to screen the drugs as inhibitors on the activity of methyltransferase in the clinic.