Genetic clocks from engineered oscillators

One defining goal of synthetic biology is the development of engineering-based approaches that enable the construction of gene-regulatory networks according to design specs generated from computational modeling. This has resulted in the construction of several fundamental gene circuits, such as toggle switches and oscillators, which have been applied in novel contexts such as triggered biolm development and cellular population control. In this talk, I will first describe an engineered genetic oscillator in Escherichia coli that is fast, robust, and persistent , with tunable oscillatory periods as fast as 13 minutes. This oscillator was designed using a previously modeled network architecture comprising linked positive and negative feedback loops. Experiments show remarkable robustness and persistence of oscillations in the designed circuit; almost every cell exhibited large-amplitude fluorescence oscillations throughout observation runs. The period of oscillation can be tuned by altering inducer levels. Computational modeling reveals that the key design principle for constructing a robust oscilla-tor is a small time delay in the negative feedback loop, which can mechanisti-cally arise from the cascade of cellular processes involved in forming a functional transcription factor. I will then describe an engineered network with global in-tercellular coupling that is capable of generating synchronized oscillations in a growing population of cells. The network is based on the interaction of two quorum sensing genes; luxI, which produces an intercellular transcriptional activator (AHL, acyl-homoserine lactone), and aiiA, which degrades AHL intracellularly. Microfluidic devices tailored for cellular populations at differing length scales are used to demonstrate collective synchronization properties along with spatiotem-poral waves occurring on millimeter scales. The period of the bulk oscillations ranges from 55-90 minutes, depending on the effective degradation rate of the AHL coupling molecule. In large monolayer colonies of cells, the time scale for the diffusive coupling of AHL is characterized by wavefront velocities that range from 8-30 microns/min.