A general approach for rational design of fluorescent DNA aptazyme sensors based on target-induced unfolding of DNA hairpins

•We developed a design of fluorescent DNA aptazyme sensors based on the target-induced unfolding of DNA hairpins.•Our approach required no change in the active sites of any DNAzyme or aptamer.•The new aptazyme sensors showed high selectivity and sensitivity for the detection of their targets.•The aptazyme sensor for interferon-gamma was the first example of an aptazyme to it.

DNA aptazymes are allosteric DNAzymes activated by the targets of DNA aptamers. They take the advantages of both aptamers and DNAzymes, which can recognize specific targets with high selectivity and catalyze multiple-turnover reactions for signal amplification, respectively, and have shown their great promise in many analytical applications. So far, however, the available examples of DNA aptazyme sensors are still limited in utilizing only several DNAzymes and DNA aptamers, most likely due to the lack of a general and simple approach for rational design. Herein, we have developed such a general approach for designing fluorescent DNA aptazyme sensors. In this approach, aptamers and DNAzymes are connected at the ends to avoid any change in their original sequences, therefore enabling the general use of different aptamers and DNAzymes in the design. Upon activation of the aptazymes by the targets of interest, the rate of fluorescence enhancement via the cleavage of a dually labeled substrate by the active aptazymes is then monitored for target quantification. Two DNAzymes and two aptamers are used as examples for the design of three fluorescent aptazyme sensors, and they all show high selectivity and sensitivity for the detection of their targets. More DNA aptazyme sensors for a broader range of targets could be developed by this general approach as long as suitable DNAzymes and aptamers are used.

Graphical abstractA simple and general approach for rational design of fluorescent DNA aptazyme sensors based on the target-induced unfolding of DNA hairpins.Figure optionsDownload full-size imageDownload as PowerPoint slide