Enzyme-free dual-amplification strategy for the rapid, single-step detection of nucleic acids based on hybridization chain reaction initiated entropy-driven circuit reaction

Developing robust, isothermal and simple nucleic acids detection strategies is of great significance to clinical diagnosis and bioanalysis. Recently, hybridization chain reaction has attracted intense interest owing to its isothermal reaction conditions, enzyme-free nature, and simple sequence design. However, strategies based on HCR alone may not offer satisfactory sensitivity when trace targets are required to be analyzed. Here, we developed an enzyme-free and isothermal dual-amplification sensing strategy for the rapid and one-step detection of nucleic acids based on hybridization chain reaction and entropy-driven circuit reaction (HCR-EDCR). The target DNA initiates upstream HCR to assemble polymeric double stranded DNA (dsDNA) nanowires composed of many tandem trigger units that motivate downstream EDCR to continuously liberate report strand, resulting in the generation of an amplified fluorescence readout signal. This isothermal and homogenous strategy exhibits high sensitivity and good selectivity with a limit of detection of 87 fM, without the involvement of any enzymes. The whole dual-amplification can be completed with one-step in 45 min. Moreover, the developed HCR-EDCR approach can be easily adapted for detecting different analytes just by substituting the target-specific sequence. Therefore, this cascaded strategy may offer a new promising model for various diagnostic and biological analysis.