Multifunctional Clip Strand for the Regulation of DNA Strand Displacement and Construction of Complex DNA Nanodevices

Abstract

Strand displacement reactions are important bricks for the construction of various DNA nanodevices, among which the toehold-mediated strand displacement reaction is the most prevalently adopted. However, only a limited number of tools could be used to finely regulate the toehold reaction, thus restricting DNA nanodevices from being more multifunctional and powerful. Herein, we developed a regulation tool, Clip, and achieved multiple regulatory functions, including subtle adjustment of the reaction rates, allosteric strand displacement, selective activation, and resetting of the reaction. Taking advantages of the multiple functions, we constructed Clip-toehold-based DNA walking machines. They showed behaviors of controllable walking, concatenation, and programmable pathways. Furthermore, we built Clip-toehold-based AND and OR logic gates and integrated those logic gates to construct multilayer circuits, which could be reset and reused to process different input signals. We believe that the proposed Clip tool has expanded the functionality of DNA strand displacement-based nanodevices to a much more complex and diverse level and anticipate that the tool will be widely adopted in DNA nanotechnology.

Cooperative Branch Migration: A Mechanism for Flexible Control of DNA Strand Displacement

“…Alternatively, Chen developed the associative toehold activation strategy that attached a DNA toehold to a BM domain through duplex formation during the execution of DNA circuits . Many other regulatory tools have also been developed, such as allosteric toehold, clip toehold, handhold, and so on. − However, toward the goal of constructing functional DNA devices of higher complexity, multiple types of regulation should be integrated to design cascades of toehold exchange reactions. Nevertheless, the majority of regulatory tools either promise a limited number of functions or are not compatible with existing DNA devices built on the conventional toehold reaction.…”

“…Nevertheless, the majority of regulatory tools either promise a limited number of functions or are not compatible with existing DNA devices built on the conventional toehold reaction. For instance, the allosteric toehold can manually control the activation of the strand displacement reaction but the assignments of the invading strand and the dissociative strand were reversed during the reaction, rendering it not compatible with most DNA devices . The clip toehold technique achieved multiple regulatory functions but required prolonged operation time to accomplish each type of regulation; thus, it is not suitable for DNA machines that require a timely operation.…”

“…For instance, the allosteric toehold can manually control the activation of the strand displacement reaction but the assignments of the invading strand and the dissociative strand were reversed during the reaction, rendering it not compatible with most DNA devices. 22 The clip toehold technique achieved multiple regulatory functions but required prolonged operation time to accomplish each type of regulation; thus, it is not suitable for DNA machines that require a timely operation. Therefore, it is worthwhile to develop regulatory tools to simplify the design of complicated strand displacement systems.…”

Constructing DNA logic circuits based on the toehold preemption mechanism

“… 24–26 The reaction rate of DNA strand displacement increases exponentially with the bonding strength, which can be increased by about 10 6 times. 27 Subsequently, in order to fine regulate the reaction rate of DNA strand displacement, a series of reaction regulation methods of toehold-mediated DNA strand displacement have been proposed, such as remote toehold, 27,28 mismatch toehold, 28,29 combination toehold, 30 and allosteric toehold. 31 These methods improve the flexibility of the toehold-mediated strand displacement reaction and provide a powerful tool for the development of DNA computing.…”

Enzymatic Cycle-Inspired Dynamic Biosensors Affording No False-Positive Identification

“…Allosteric regulation of biomolecules and chemical events is an intriguing natural phenomenon. Its significance is not just crucial for dynamic control of biological processes, but it also inspires toolboxes for programmable synthetic nanodevice engineering. − Over the past two decades, there has been considerable progress in the design of controllable biosensor components, − precision switches, − and genetic regulators − based on the principle that the activity of the synthetic structure corresponds to a varied free-energy state regulated by an allosteric effector. These efforts have focused on exploring new allosteric toolboxes to construct synthetic allosteric systems. − However, designing synthetic allosteric systems that can reversibly switch their conformational states is more challenging because multiple states relative to positive/negative activities must be flexible enough that switching can be reversibly toggled by allosteric effectors.…”