The detection of nucleic acids is fundamental for studying their functions and for the development of biological studies and medical diagnostics. Herein, we report a new strategy for nucleic acid amplified detection by combining target-catalyzed dynamic assembly with host-guest interaction between β-cyclodextrin polymer (β-CDP) and pyrene. In this strategy, a metastable pyrene-labeled hairpin DNA probe (probe H1) and a metastable unlabeled hairpin DNA probe (probe H2) were elaborately designed as the assembly components, which were kinetically handicapped from cross-opening in the absence of target DNA. In this state, pyrene labled at the 5'-termini of single-stranded stem of probe H1 would be easily trapped into the hydrophobic cavity of β-CDP because of weak steric hindrance, leading to significant fluorescence enhancement. Once the dynamic assembly was catalyzed by target DNA, a hybridized DNA duplex H1-H2 would be created continuously. In this state, it is difficult for pyrene to enter the cavity of β-CDP due to steric hindrance and weak-binding interaction, leading to a weak fluorescent signal. Thus, target DNA could be detected by this simple mix-and-detect amplification method without the need of expensive and perishable protein enzymes. As low as 10 pM of the target DNA was detected by this assay, which was comparable to that of some reported enzyme-dependent amplification methods. Meanwhile, the proposed method was further successfully applied to detect DNA in cell lysate samples, showing great potential for target detection from complex fluids. In addition, as a novel transformation of dynamic DNA assembly technology into enzyme-free signal-amplification analytical application, the proposed strategy has shown great potential for applications in a wide range of fields, such as aptamer-based non-nucleic acid target sensing, biomedicine and bioimaging.
Keywords: Enzyme-free; Nucleic acid detection; Pyrene; β-cyclodextrin polymer.
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