Encoding DNA Frameworks for Amplified Multiplexed Imaging of Intracellular microRNAs

Real-time imaging of multiple low-abundance microRNAs (miRNAs) simultaneously in living cells with high sensitivity is of vital importance for accurate cancer clinical diagnosis and prognosis studies. Maintaining stability of nanoprobes resistant to enzyme degradation and enabling effective signal amplification is highly needed for in vivo imaging studies. Herein, a rationally designed one-pot assembled multicolor tetrahedral DNA frameworks (TDFs) by encoding multicomponent nucleic acid enzymes (MNAzymes) was developed for signal-amplified multiple miRNAs imaging in living cells with high sensitivity and selectivity. TDFs could enter cells via self-delivery with good biocompatibility and stability. Two kinds of MNAzymes specific for miRNA-21 and miRNA-155 with fluorescein labeling were encoded in the structure of TDFs respectively through one-step thermal annealing. In the intracellular environment, the TDFs could be specifically bound with its specific miRNA target and form an active DNAzyme structure. The cleavage of the active site would trigger the release of target miRNA and circular fluorescence signal amplification, which enabled accurate diagnosis on miRNA identifications of different cell lines with high sensitivity. Meanwhile, with the specific AS1411 aptamer targeting for nucleolin overexpressed on the surface of the carcinoma cells, this well-designed TDFs nanoprobe exhibited good discrimination between cancer cells and normal cells. The strategy provides an efficient tool for understanding the biological function of miRNAs in cancer pathogenesis and therapeutic applications.