Molecular Computers Built from DNA Components

Thesis (Ed.D.)--University of Washington, 2020At the nanoscale, the ability to control spatio-temporal interactions would give us unparalleled power. DNA strand displacement systems seem to be the perfect technology to achieve control over molecular interactions, as they have the major advantage that reactions can be predicted from domain structure alone. Strand displacement technology has resulted in a myriad of dynamic devices with applications for everything from diagnostics to biomimetic manufacturing. Our goal is to build DNA strand displacement computers to achieve control over the temporal dynamics of molecules, meaning how they interact in time, and also the spatial dynamics, or how they interact in space. To build these types of molecular computers, we take inspiration from the programmability of silicon computers, which take programming languages as input. As a primitive for a molecular programming language, we look to previous work which has shown that the behavior of formal chemical reaction networks (CRNs) can be approximated using nicked double stranded DNA (ndsDNA) gates. CRNs are a natural way to think about molecular interactions and have been shown to be Turing complete, if used as a programming language. To compose complex or large DNA strand displacement CRNs, it is desirable to compose them from small, well-characterized systems, a strategy that requires quantitatively predictable reaction kinetics. However, parameter estimation of these ndsDNA gates has thus far required fitting reaction rates for every strand displacement occurrence individually and these fitted reaction rates were found to vary over more than an order of magnitude despite toehold sequences designed to have the same length and GC content. With our work on context-independent plasmid-derived gates, high quality fits can be obtained using a single reaction rate constant k for all strand displacement steps, allowing for predictable and composable kinetic behavior. Additionally, the nicked double stranded structure of the gates allows for them to be derived from plasmids as a source for highly pure DNA for use in strand displacement experiments. In our work, we use a cloning strategy that dramatically reduces recombination events, increasing the yield of functional gates that can be used for experiments, and can also be used to control stoichiometry of chemical reactions. Here, we also show that our platform using context-independent plasmid-derived gates can also be used in a spatial setting. We built a travelling wave system by putting a synthetic autocatalytic reaction in a spatial setting, demonstrating the first steps towards an autonomous and synthetic pattern formation system on a centimeter scale based on a DNA strand displacement system