A novel DNA biosensor was fabricated with three-dimensionally ordered macroporous gold-nanoparticle-doped titanium dioxide(3DOM GTD) film on the indium-tin oxide(ITO) electrode surface by the colloidal crystal template technique,and calf thymus DNA(ctDNA)was immobilized on the 3DOM GTD/ITO successfully.The morphology of the modified electrode was characterized by transmission electron microscopy(TEM) and scanning electron microscopy(SEM).The immobilization of ctDNA on the surface of 3DOM GTD/ITO modified electrode was investigated by electrochemical impedance spectroscopy(EIS).The EIS experimental results showed that ctDNA was successfully immobilized on the 3DOM GTD/ITO modified electrode.The electrochemical properties of the antitumor drug quercetin (Qu) and the interaction between Qu and ctDNA were studied on 3DOM GTD/ITO modified electrode by cyclic voltammograms (CV) and differential pulse voltammograms(DPV) method.The result showed that Qu had a pair of quasi-reversible redox peak on 3DOM GTD/ITO modified electrode,and the transfer process of 2 electrons and 2 protons was an involved in the electrode reaction.There was a strong interaction between Qu and ctDNA immobilized on the modified electrode,and the binding constant K was 3.61×106L/mol.The binding mode between Qu and ctDNA was an intercalation confirmed by CV,UV-Vis absorption spectra (UV-Vis),fluorescence spectra(FL) and circular dichroism spectra(CD) experiments. The sequence selectivity of binding between Qu and ctDNA was investigated,and the binding constants of Qu and poly(dG-dC) and poly(dA-dT) were calculated, respectively, according to the results of CV experiments.The binding constant ratio K(dG-dC)/K(dA-dT) was obtained to be 3.5,indicating that Qu bound to the GC region preferentially when Qu interacted with ctDNA.