Label-Free Detection of Cu2+ and Hg2+ Ions Using Reconstructed Cu2+-Specific DNAzyme and G-quadruplex DNAzyme

Abstract

Label-free metal ion detection methods were developed. To achieve these, a reconstructed Cu2+-specific DNA-cleaving DNAzyme (Cu2+-specific DNAzyme) with an intramolecular stem-loop structure was used. G-quadruplex-forming G-rich sequence(s), linked at the ends of double-helix stem of an intramolecular stem-loop structure, was partly caged in an intramolecular duplex or formed a split G-quadruplex. Cu2+-triggered DNA cleavage at a specific site decreased the stability of the double-helix stem, resulting in the formation or destruction of G-quadruplex DNAzyme that can effectively catalyze the 2,2′-azinobis(3-ethylbenzothiazoline)-6-sulfonic acid (ABTS)-H2O2 reaction. Based on these, two label-free, cost-effective and simple Cu2+ sensors were designed. These two sensors followed different detection modes: 'turn-on' and 'turn-off'. As for the 'turn-on' sensor, the intramolecular stem-loop structure ensured a low background signal, and the co-amplification of detection signal by dual DNAzymes (Cu2+-specific DNAzyme and G-quadruplex DNAzyme) provided a high sensitivity. This sensor enabled the selective detection of aqueous Cu2+ with a detection limit of 3.9 nM. Visual detection was possible. Although the 'turn-off' sensor gave lower detection sensitivity than the 'turn-on' one, the characteristics of cost-effectiveness and ease of operation made it an important implement to reduce the possibility of pseudo-positive or pseudo-negative results. Combining the ability of Hg2+ ion to stabilize T-T base mismatch, above dual DNAzymes-based strategy was further used for Hg2+ sensor design. The proposed sensor allowed the specific detection of Hg2+ ion with a detection of 4.8 nM. Visual detection was also possible. © 2013 Li et al.