Abstract: In this study, various types of self-complementary secondary structures were introduced into the input single strand DNA of strand displacement reactions to change their free energy and conformations, aiming to investigate the influence of different structures on the rate of strand displacement reaction. We found that the reaction rate decreased as the length of the self-complementary structure increased. This effect was more pronounced when the self-complementary structure was designed in the toehold domain than when it formed in the non-toehold region. The incorporation of secondary structures into the input strands led to varying degrees of reduction in strand displacement reaction rates, and in some cases, completely inhibited the reaction. Finally, we elucidate the mechanism by which secondary structures influence the kinetics of strand displacement reactions, offering new insights into expanding the temporal regulation capabilities of DNA strand displacement and advancing its applications in DNA molecular computing and biosensing.

Key words: DNA strand displacement reaction, Secondary structure, Reaction kinetics