Mismatches Improve the Performance of Strand‐Displacement Nucleic Acid Circuits

Abstract Catalytic hairpin assembly (CHA) has previously proven useful as a transduction and amplification method for nucleic acid detection. However, the two hairpin substrates in a CHA circuit can potentially react non‐specifically even in the absence of a single‐stranded catalyst, and this non‐specific background degrades the signal‐to‐noise ratio. The introduction of mismatched base pairs that impede uncatalyzed strand exchange reactions led to a significant decrease of the background signal, while only partially damping the signal in the presence of a catalyst. Various types and lengths of mismatches were assayed by fluorimetry, and in many instances, our MismatCHA designs yielded 100‐fold increased signal‐to‐background ratios compared to a ratio of 4:1 with the perfectly matched substrates. These observations could be of general utility for the design of non‐enzymatic nucleic acid circuits.