Microfluidic platforms for the engineering of continuously-operating, synthetic nucleic acid-based systems

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Nucleic acid nanotechnology is a versatile platform for engineering molecular networks. By designing oligonucleotide molecules with specific sequences, one can build complex synthetic systems with predictable and programmable reactions. The most versatile networks are based on "strand displacement" reactions, in which base pairing is used to drive the replacement of one or more strands in a multi-stranded "gate complex". The outputs of these displacement reactions can trigger subsequent reactions, allowing the construction of large networks.Traditionally, the essential gate complexes are produced via a multi-step process that cannot be realised in situ. Individual strands are separately synthesized, annealed to form multi-stranded gates, and these gates are then mixed to create a network that produces a single fixed output. Traditional strand displacement-based networks cannot operate continuously without an external supply of gates produced in this way. This limitation prohibits their application in engineered or synthetic cells, where networks must continuously respond to their environment using components produced in situ and in real time.We have recently demonstrated in situ production of multi-stranded RNA gate molecules directly from transcription, utilising self-cleaving RNA ribozyme motifs that convert a folded single-stranded RNA transcript into a multi-stranded complex [Bae et al., https://pubs.acs.org/doi/10.1021/acs.nanolett.0c03629]. These constructs have the potential to underlie continuously-operating strand displacement networks; this project aims to explore this potential using microfluidic "cells" that can sustain continuously-operating networks. The result will be a novel engineering platform for nucleic acid nanotechnologists, and also a vital step in the process of incorporating nucleic acid-based circuits into living cells.The student will start by developing microfluidic platforms that are appropriate to hosting continuously active nucleic acid-based systems, identifying optimal geometries and materials for the challenge. Subsequently, the student will test nucleic acid-based circuits of increasing complexity in the microfluidic chips built. Both stages of the project will involve significant input from computational modelling; the project is therefore highly appropriate for students seeking an interdisciplinary challenge straddling engineering, nanotechnology and biology.