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  • Title: Construction of an enzyme-free concatenated DNA circuit for signal amplification and intracellular imaging.
    Author: Wang H, Li C, Liu X, Zhou X, Wang F.
    Journal: Chem Sci; 2018 Jul 14; 9(26):5842-5849. PubMed ID: 30079197.
    Abstract:
    Nucleic acid circuits have shown promising potential for amplified detection of biomarkers with interest in biologically important engineering applications. In this work, by properly integrating two signal amplification approaches, catalytic hairpin assembly (CHA) and hybridization chain reaction (HCR), a concatenated CHA-HCR system was established as an isothermal enzyme-free amplification strategy for highly sensitive and selective nucleic acid assay. The target catalyzes the self-assembly of CHA hairpin substrates into dsDNA products, where the split segments of HCR trigger are successively connected to drive the subsequent autonomous cross-opening of HCR hairpins, leading to the construction of HCR tandem copolymeric dsDNA nanowires. The resulting HCR copolymer brings a fluorophore donor/acceptor pair into close proximity that allows an efficient generation of FRET readout signal. Moreover, the optimized CHA-HCR circuit, upon the incorporation of an auxiliary sensing module, can be converted into a universal sensing platform for detecting cancerous biomarkers (e.g., a well-known oncogene miR-21) through a convenient easy-to-integrate procedure. The concatenated CHA-HCR amplifier enables accurate intracellular miRNA imaging in living cells, which is especially suitable for in situ amplified detection of lowly expressed endogenous analytes. The inherent synergistically accelerated recognition and hybridization features of CHA-HCR circuit contribute to the amplified detection of endogenous RNAs in living cells. The flexible and programmable nature of the homogeneous CHA-HCR system provides a versatile and robust toolbox for a wide range of research fields, such as in vivo bioimaging, clinical diagnosis and environmental monitoring.
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