The objective of this project is to generate robust and tunable molecular clocks using nucleic acids and proteins. Oscillators are fundamental components both in biology and in computation: molecular clocks synchronize a variety of cellular processes, as much as computer clocks orchestrate the operations of millions of silicon devices. This research will combine dynamical systems tools and recent advances in nucleic acids nanotechnology to systematically study the design and synthesis of biochemical clocks in vitro. An array of reaction primitives will be generated for dynamic production, degradation and transformation of nucleic acids and protein components. These reaction primitives will be used to build candidate clocks, whose robustness will be evaluated in silico using structural methods (graphical and geometric conditions) and numerical bifurcation analysis. In particular, research will focus on creating clocks with separately tunable frequency and amplitude. The best candidate oscillators will be synthesized by specifying nucleic acid sequences of the primitives with available software tools. Experimental screening will be performed with a high-throughput, randomized experimental setup relying on microdroplets.
This research will expand scientific knowledge regarding the structure of robust and tunable oscillators. In particular, this project will bridge the gap between theoretical studies aimed at discovering periodic behaviors in biochemical networks and their systematic experimental implementation. This bridge can be built because of the programmability of nucleic acid networks. Our results will have an impact in two main areas: 1) New bottom-up design principles will be validated integrating dynamical systems theory and high-throughput experimental techniques. This platform will be used to generate other autonomous biochemical circuits, such as multistationary systems and complex nonlinear networks; 2) The availability of programmable timing devices will enable the coordination of logic circuits, molecular machines, reconfigurable nanostructures, and pattern formation. The generated clocks will be useful in a variety of contexts including nanofabrication, biomaterials, and drug development, because nucleic acids can be selectively interfaced with a variety of organic and inorganic materials. In addition, theoretical and computational results will provide novel criteria to investigate the features of natural oscillators, and will open new avenues to the use of programmable nucleic acid circuits for generating periodic behaviors in cells. The results of this research will be disseminated through published materials and international conference presentations. Interdisciplinary training will be provided to participating graduate and undergraduate students. The PI's group will pursue outreach activities targeting K8-12 students at local underserved schools, including the creation of mini-courses and videos about periodic phenomena and timekeeping in technology and biology.
