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Team:ETH Zurich/achievements
From 2013.igem.org
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<h1> We achieved </h1> | <h1> We achieved </h1> | ||
<b>Pre-Processing:</b><br><br> | <b>Pre-Processing:</b><br><br> | ||
| - | - 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 LuxI promoter and the incubation time for the diffusion.<br> | + | - 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 LuxI promoter and the incubation time for the diffusion.<br> |
- 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.<br> | - 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.<br> | ||
- Make a spatio-temporal model of AHL difussion<br><br> | - Make a spatio-temporal model of AHL difussion<br><br> | ||
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- To make a promoter libary by partial sensitivity recovery based on the first PLuxR variant and finally isolate 8 promoters with different EC<sub>50</sub>.<br> | - To make a promoter libary by partial sensitivity recovery based on the first PLuxR variant and finally isolate 8 promoters with different EC<sub>50</sub>.<br> | ||
- We measured the dose-response of those promoters in liquid culture as well as on agar plates and determined the EC<sub>50</sub> going from 0.02nM to 6482nM in liquid culture and from 4.45nM to 12'555nM on agar plates.<br> | - We measured the dose-response of those promoters in liquid culture as well as on agar plates and determined the EC<sub>50</sub> going from 0.02nM to 6482nM in liquid culture and from 4.45nM to 12'555nM on agar plates.<br> | ||
| - | - By the prediction of the model we chooose xy to be the right one for our set-up.<br> | + | - By the prediction of the model we chooose xy to be the right one for our set-up.<br><br> |
<i> A proof-of-principle using a GFP reporter.</i><br><br> | <i> A proof-of-principle using a GFP reporter.</i><br><br> | ||
Revision as of 09:14, 27 October 2013
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 LuxI promoter 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 PLuxR 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:
- Identification of the leakyness source.
- Optimize the circuit to reduce the leakiness by introduction of a negative feedback loop.
Output:
- Implement several different reporters (hydrolases) in one construct.
- Different colormetric response for all hydrolases.
- 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.
