SKM 2023 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 11: Poster Session I

BP 11.72: Poster

Dienstag, 28. März 2023, 12:30–15:30, P1

Steady-state operation of a cell-free genetic band-detection circuit — •Anna C. Jäkel, Lukas Aufinger, and Friedrich C. Simmel — Technichal University of Munich, Munich, Germany

Synthetic gene networks have been used extensively to explore principles of biological pattern formation as they play a decisive role during biological growth and development processes. Here, we report on a bottom-up approach to design and analyze a cell-free genetic circuit based on an incoherent feed forward loop (IFFL-2), which is expected to produce a three-stripe pattern in response to an input gradient. In our work, we first simulated the behavior of the circuit and explored relevant parameters using a genetic algorithm. We then tested our circuit in a bacterial cell-free gene expression system and found that our circuit is only functional under non-equilibrium conditions in microfluidic ring reactors, whereas it fails to perform in bulk experiments in closed reactors. Hence, we concluded that non-equilibrium conditions are of necessity to establish the double-repression cascade which was the essential element of the genetic circuit. We used six neighboring ring reactors to establish a 'virtual' morphogen gradient by supplying the reactors with decreasing amounts of the transcription factor sigma28, corresponding to the different positions within an exponential morphogen gradient. We finally demonstrated that our IFFL-2 circuit, when operated in the microfluidic system, shows the correct gene expression response that is required for stripe-formation in a spatial context.