We achieved

Pre-Processing:

- Characterize the AHL diffusion on agar plates and define, with the predition from the model and the experimental results, the distance between colonies, the strengh of the promoter controlling LuxI production and the incubation time for the diffusion.
- To astablish several AHL gradients from different mines on one plate, intersections of gradients result in higher AHL levels for the detection of 1, 2 or 3 mines.
- Make a spatio-temporal model of AHL difussion

Processing:

- To isolate a first P_LuxR by site directed mutagenisis.
- Find a analytical solution for the EC50 of the promoter we need to find.
- To make a promoter libary by partial sensitivity recovery based on the first PLuxR variant and finally isolate 8 promoters with different EC₅₀.
- We measured the dose-response of those promoters in liquid culture as well as on agar plates and determined the EC₅₀ going from 0.02nM to 6482nM in liquid culture and from 4.45nM to 12'555nM on agar plates.
- By the prediction of the model we chooose xy to be the right one for our set-up.

- a proof-of-principle using a GFP reporter.

Optimization:

- the identification of the leakyness source.
- to model different solution to reduce the leakiness - to optimize the circuit to reduce the leakiness by introduction of a negative feedback loop.

Output:

- to implement several different reporters (hydrolases and fluorencent proteins) in one construct.
- to show different colormetric (yellow, salmon, violett, green, blue) response for 5 hydrolases.
- to show the orthogonality of all 5 hydrolases.
- to characterize the hydrolases by Michaelis-Menten kinetics.


A preliminary game with the hydrolases.

Establish a spatio-temporal model of the proof-of-principle and the final game.

We thank our sponsors: