Spatiotemporal Monitoring of Subcellular mRNAs In Situ via Polyadenine-Mediated Dual-Color Sticky Flares.

Abstract

Spatiotemporal monitoring of multiple low-abundance messenger RNAs (mRNAs) is vitally important for the diagnosis and pathologic analysis of cancer. However, it remains a clinical challenge to monitor and track multiple mRNAs location simultaneously in situ at subcellular level with high efficiency. Herein, we proposed polyA-mediated dual-color sticky flares for simultaneous imaging of two kinds of intracellular mRNA biomarkers. Two kinds of fluorescent DNA specific for GalNac-T mRNA and c-Myc mRNA were functionalized onto gold nanoparticles (AuNPs) through efficient polyadenine (polyA) attachment. By tuning polyA length, the lateral spacing and densities of DNA on AuNPs could be precisely engineered. Compared to the traditional thio-DNA-modified nanoprobes, the uniformity, detection sensitivity, and response kinetics of sticky flares were greatly improved, which enables live-cell imaging of mRNAs with enhanced efficiency. With a sticky-end design, the fluorescent DNA could dynamically trace mRNAs after binding with target mRNAs, which realized spatiotemporal monitoring of subcellular mRNAs in situ. Compared to one target mRNA imaging mode, the multiple target imaging mode allows more accurate diagnosis of cancer. Furthermore, the proposed polyA-mediated dual-color sticky flares exhibit excellent cell entry efficiency and low cytotoxicity with a low-cost and simple assembling process, which provide a pivotal tool for multiple targets imaging in living cells.