One-step self-assembly of quantum dot-based spherical nucleic acid nanostructure for accurate monitoring of long noncoding RNA MALAT1 in living cells and tissues

Simple and sensitive measurement of long non-coding RNA (lncRNA) is critical for early detection of malignancies. Herein, we demonstrate one-step self-assembly of quantum dot (QD)-based spherical nucleic acid (SNA) nanostructure for accurate monitoring of lncRNAs in living cells and tissues. When target lncRNA is present, it binds with a dumbbell probe to expose the complementary domain of Cy5-labeled primer, which subsequently induces cascade primer exchange reaction to produce abundant Cy5-labeled initiators. The Cy5-labeled initiators subsequently hybridize with hairpin probes on the QD surface to activate isothermal circular strand-displacement polymerization reaction, generating the QD-DNA-Cy5 nanostructures and inducing efficient Förster resonance energy transfer (FRET) between donor QD and acceptor Cy5. The obtained FRET signals are accurately quantified by single-molecule imaging. Notably, the single QD-based SNA nanostructure functions not only as a signal transmitter but also as a protector against non-specific amplification. Moreover, this assay utilizes only one DNA polymerase to achieve two-stage amplification, avoiding careful modulation of multiple enzymes. The self-assembly of QD nanosensor can be accomplished in single-step and single-tube manners at room temperature, eliminating precise temperature control and labor-intensive reaction protocols. This QD nanosensor achieves high sensitivity with a limit of detection (LOD) of 65.25 aM, and it is capable of quantifying lncRNA expression at single-cell level, differentiate tumor cells from normal cells, and distinguish breast cancer patients from healthy individuals, providing a versatile paradigm for biomedical research and early clinic diagnostics.