Adverse Effects of UV Exposure on DNA Strand Displacement Reactions

DNA strand displacement (DSD) reactions are widely used in molecular computing and nanotechnology due to their programmability and precise control over molecular interactions. However, experimental DSD systems often underperform relative to theoretical models, due in part to poorly characterized sources of reactant impurities. Here, we identify UV shadowing (a standard technique for visualizing DNA during PAGE purification) as a previously overlooked cause of DSD circuit errors. Counterintuitively, existing purification protocols with UV shadowing can increase impurities that disrupt circuit behavior. Specifically, we demonstrate that UV exposure (i) reduces DSD reaction percent yield and (ii) leads to a phenomenon we call "negative leak," i.e., the attenuation of the free functional signal due to its sequestration on double-stranded complexes that are designed to be inert. Additionally, we quantify the sequence dependence of UV-induced DSD errors, which increase with the number of adjacent pyrimidine pairs. To circumvent these errors, we introduce a simple and practical purification protocol that avoids UV exposure and show that the errors no longer occur. Our results highlight UV-induced damage as a critical factor in DSD circuit performance and suggest that UV shadowing may have contributed to significant reproducibility and scaling challenges in the broader DSD literature.