Exceptional quenching properties of tetrazine-based organic frameworks for fluorescently labeled nucleic acids and their applications in sensing

Covalent organic frameworks (COFs) have garnered significant attention due to their unique properties, such as high surface area, porosity, chemical stability, and sustainability, which enable a wide range of applications in recent years. In this study, tetrazine-based organic frameworks, named TzF-9, were investigated for their ability to quench fluorescence in nucleic acids labeled with fluorophores. The experimental results demonstrated that TzF-9 effectively quenched the fluorescence of fluorophore-labeled single-stranded deoxyribonucleic acid (ssDNA) probes with more than six bases. Notably, the quenching process was rapid, reaching equilibrium in just about three minutes to achieve a high quenching efficiency (∼95%). Significantly, its excellent quenching ability is retained across a wide pH range. Furthermore, fluorescent dyes, including fluorescein (FAM), cyanine dye 3 (Cy3), and 6-carboxy-X-rhodamine (ROX), labeled on long ssDNA probes, were efficiently quenched, indicating that TzF-9 can function as a universal fluorescence quencher. In addition, the quenching efficiency of TzF-9 for short ssDNA and double-stranded DNA (dsDNA) probes was significantly lower than for long ssDNA probes. Taking advantage of these distinct quenching efficiencies for DNA probes with different structures, TzF-9 was employed as a sensing platform for detecting ssDNA and nuclease activity, exhibiting good selectivity and high sensitivity. With its combination of strong quenching ability and high stability, TzF-9 presents a promising quencher for biosensing applications.