Sustainable and cascaded catalytic hairpin assembly for amplified sensing of microRNA biomarkers in living cells

The sensing of intracellular microRNAs (miRNAs) is of significance for early-stage disease diagnosis and therapeutic monitoring. DNA is an interesting building material that can be programed into assemblies with rigid and branched structures, especially suitable for imaging intracellular biomolecules or therapeutic drug delivery. Here, by introducing the palindromic sequences into the programmable DNA hairpins, we describe an endogenous target-responsive three-way branched and palindrome-assisted catalytic hairpin assembly (3W-pCHA) approach for imaging miRNA-155 of living tumor cells with high sensitivity. The miRNA-155 triggers autonomous assembly of the fluorescently quenched signal hairpin and two hairpin dimers formed via hybridization of their respective palindromic sequences to yield branched DNA junctions, which carry the unopened hairpins and thus provide addressable substrates for continuous assembly formation of DNA nanostructures. During the formation of the DNA nanostructures, the miRNA-155 is cyclically reused and many signal probes are unfolded to show highly intensified fluorescence for detecting miRNA-155 down to 6.9 pM in vitro with high selectivity. More importantly, these probes can be transfected into live cancer cells to initiate the assembly process triggered by intracellular miRNA-155, which provides a new way for imaging highly under-expressed miRNAs in cells. Besides, this approach can also be employed to differentiate miRNA-155 expression variations in different cells, indicating its promising potentials for early-stage disease diagnosis and biological studies in cells.