DNA Tetrahedra Modules as Versatile Optical Sensing Platforms for Multiplexed Analysis of miRNAs, Endonucleases, and Aptamer-Ligand Complexes.

Zhixin Zhou,Yang Sung Sohn,Rachel Nechushtai,Itamar Willner

doi:10.1021/acsnano.0c04031

Abstract

The sensing modules for analyzing miRNAs or the endonucleases consist of tetrahedra functionalized with three different fluorophore–quencher pairs in spatially quenched configurations and hairpin units acting as recognition elements for the analytes. Three different miRNAs (miRNA-21, miRNA-221, and miRNA-155) or three different endonucleases (Nt.BbvCI, EcoRI, and HindIII) uncage the respective hairpins, leading to the switched-on fluorescence of the respective fluorophores and to the multiplex detection of the respective analytes. In addition, a tetrahedron module for the multiplexed analysis of aptamer ligand complexes (ligands = ATP, thrombin, VEGF) is introduced. The module includes edges modified with three spatially separated fluorophore–quencher pairs that were stretched by the respective aptamer strands to yield a switched-on fluorescent state. Formation of the respective aptamer ligands reconfigures the edges into fluorophore-quenched caged-hairpin structures that enable the multiplexed analysis of the aptamer–ligand complexes. The facile permeation of the tetrahedra structures into cells is used for the imaging of MCF-7 and HepG2 cancer cells and their discrimination from normal epithelial MCF-10A breast cells.

Highlights

The sensing modules for analyzing miRNAs or the endonucleases consist of tetrahedra functionalized with three different fluorophore−quencher pairs in spatially quenched configurations and hairpin units acting as recognition elements for the analytes
Hybrid conjugates consisting of hemin/G-quadruplex and different-sized semiconductor quantum dots were applied for the multiplexed analysis of genes[16] and aptamer−ligand complexes[17] using chemiluminescence resonance energy transfer (CRET) as the readout signal
We describe the engineering of the edges of the tetrahedra with three different fluorophore−quencher pairs and with predesigned hairpin structures and present the use of the tetrahedra as functional modules for the parallel and multiplexed sensing of different analytes

Summary

Introduction

The sensing modules for analyzing miRNAs or the endonucleases consist of tetrahedra functionalized with three different fluorophore−quencher pairs in spatially quenched configurations and hairpin units acting as recognition elements for the analytes. The treatment of the tetrahedron module with EcoRI leads to the cleavage of the stem domain of hairpin H5, and this transforms the H5-modified edge into a broken domain where the fluorophore ROX is spatially separated from the quencher, state VII.

Results

Conclusion