Real-time label-free analysis of the thermostability of DNA structures using GelRed

摘要

In biological systems, conformational transformations of nucleic acids play critical roles in genetic regulation. However, it remains a tricky task to design and optimize specific labeling strategies to track these changes.In this study, we exploited an intercalating fluorescent dye,GelRed, to characterize different DNA structures. We studied the correlation between fluorescence intensity and DNA structural properties. We showed that single-stranded DNAs with predicted self-folded secondary structures show much stronger fluorescence than those without such structures. For double-stranded DNAs, we observed that fluorescence intensity is positively correlated to their GCcontent. We also demonstrated that GelRed can be used to monitor DNA conformational changes upon temperature variations in real time. Based on these findings, we concluded that the fluorescence intensity of a GelRed-stained DNA structure has a good correlation with its thermostability in the form of a change in Gibbs free energy.