Adverse Effects of UV-Exposure on DNA Strand Displacement Reactions

Abstract

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," where high-concentration double-stranded complexes that are designed to be inert can sequester functional signal strands and thus disrupt circuit performance. Additionally, we find that UV damage is sequence-dependent, with adjacent pyrimidine-pair-rich strands being more susceptible to errors. 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.