Branched DNA Junction-Enhanced Isothermal Circular Strand Displacement Polymerization for Intracellular Imaging of MicroRNAs.

Abstract

The microRNA profiles within living cells are informative for diagnosis and prognosis of human cancers. In the present work, we developed a new sensing strategy based on branched DNA junction-enhanced isothermal circular strand displacement polymerization (B-ICSDP) for the detection and intracellular imaging of microRNAs in living cells of interest. A circular DNA template consisting of three repetitive fragments serves as the scaffold for the self-assembly of sophisticated signaling probes, resulting a shrunk branched DNA junction. Target microRNA triggers the opening of molecular beacon, not only restoring the quenched fluorescence but also activating a circular polymerization-based strand displacement reaction. Thus, patulous branched DNA junction is abundantly formed, generating the amplified signal. It is noteworthy that great heaps of branched product assemblies can be also achieved in living cells, and the intracellular enzymatic assembly based strategy is able to be used to recognize specific microRNA-expressed cancer cells. Moreover, different microRNAs coexisting in the same living cells can be simultaneously screened without any interference from each other by confocal laser scanning microscopy. The measured data from confocal fluorescence imaging of different cancer cells demonstrates that the B-ICSDP-based system is a promising alternative for in vivo analysis of microRNAs in complicated biological samples.