Catalytic hairpin assembly-based double-end DNAzyme cascade-feedback amplification for sensitive fluorescence detection of HIV-1 DNA

In this work, a simple all-nucleic acid cascade-feedback amplification strategy for homogeneous and protein enzyme-free fluorescence detection of HIV-1 related DNA (HIV-1 DNA) has been proposed by integrating catalytic hairpin assembly (CHA) circuit with double-end Mg2+-dependent DNAzyme autocatalytic feedback amplification. Here, the active double-end DNAzyme assemblies were derived from target-catalyzed CHA circuit, which further circularly cleaved the ribonucleotide-containing quenched fluorogenic hairpin substrates to generate distinctly amplified fluorescence signal. Meanwhile, the released quencher-labeled fragments as target DNA analogues were also able to autocatalyze CHA-DNAzyme reaction process, thus improving the determination sensitivity of HIV-1 DNA. The result demonstrated that the fluorescence intensity increment of double-end DNAzyme was over 3 times higher than that of single-end DNAzyme. The sensing method displayed a good linear range from 1 pM to 2 nM with a detectable minimum concentration of 1 pM and high specificity towards different mismatched target DNAs. Moreover, the practical application potential of the proposed method for target DNA detection in complex biological matrices was also assessed. Considering the appealing feature of programmable nucleic acids in CHA-DNAzyme sensing platform, the current strategy may provide a prospective design for detection of broad-spectrum nucleic acid biomarkers.