Formation of DNA duplexes in the presence of urea as a chaotropic agent

Hybridization of fluorescent molecular probes, such as molecular beacons or linear molecular probes, with their molecular targets can be confirmed through fluorescence spectra. In this study, we investigate the interaction between fluorescent probes, specifically linear molecular probes, and their targets in the presence of chaotropic agents. Our findings show that double-stranded structures containing mismatched nucleotides (e.g., single-nucleotide polymorphisms) occupy lower energy levels (red shifted) compared to those without mismatched nucleotides. Molecular duplexes formed in buffers without chaotropic agents do not exhibit significant differences, independently on whether they contain mismatched nucleotides or are formed with perfectly matching targets. This effect appears in the presence of a naturally occurring chaotropic agent, urea, and was confirmed in the concentration range of 1 mM to 4 mM. These findings suggest that urea and similar agents may play a significant role in the formation of mismatched nucleotide structures.