Abstract

Alfonso Jaramillo

The design and implementation of small synthetic genetic circuits for cell reprogramming is propelling the emerging field of Synthetic Biology. To facilitate their construction, libraries of standardized genetic parts are used to assemble synthetic circuits. The advent of a standardized kinetic characterization of such parts is making possible to predict their dynamical behaviour after assembly.

However, there are no automated design methods that exhaustively explore in silico all this combinatorial genetic diversity towards a circuit with targeted behaviour. We developed a model-based design platform that harnesses a library of standard genetic parts to optimize a circuit according to the desired design specifications. We have illustrated the power and versatility of our approach by designing several genetic circuits working as band-detectors, oscillators and counters. We showed that even a registry of few parts is able to contain a rich spectrum of dynamical behaviours, provided some key genetic parts are available. We have proposed a new mechanism to generate developmental patterns and oscillations using a minimal number of regulatory elements. For this, we design a synthetic gene circuit with an antagonistic self-regulation to study the spatiotemporal ontrol of protein expression. We have constructed and characterised in E. coli minimal gene networks with oscillatory behaviour. We use microfluidic techniques to track the single-cell dynamics for several days.

We have also engineered for the first time coupled oscillators in a single cell. Coupling of two oscillators is known in physics to generate a number of interesting dynamical behaviours. The resulting function could represent a simple super-position of the dynamical behaviour or it could lock into several possible characteristic frequencies, or even it could have several characteristic properties, depending on the conditions of the experiment. To analyse these effects in vivo we designed and constructed several genetic parts that allow us to characterise the dynamical behaviour of a coupled oscillator system in bacteria. Our engineered gene networks could be used in larger systems, opening the way for the engineering of genetic circuits with high complex behaviour.