Team:CIDEB-UANL Mexico/Project-Circuit

Circuit

The circuit works as the following way.
1. A pR (repressible promoter) is active and produces the Vip3Ca3 protein and the Green Fluorescent Protein as a reporter of the Vip3Ca3 production.
2. In the other part, there is a Constitutive promoter (pConst) but it works with a riboswitch.
3. The riboswitch is activated after the 32ºC and it starts trasnlation producing the cI protein which inhibits the pR turning off the Vip3Ca3 production.

One system is going to be repressing the other one, with the riboswitch when the specific range of temperature is available, from 20oC to 37oC, but when the temperature is lower to this interval the first system is going to be off and the second system is going to be active with the production of the pesticide (Vip3Ca3) and GFP as a reporter. When the temperature is low, the promoter (J23100) is going to be inhibited and then the first system it’s going to be shut down. The second system is activated and the production of the pesticide (Vip3Ca3) is on, as well as GFP production as a sign of production. As we know, the part that has the GFP reporter will be repressed after the 37ºC so it needs a degradation tag to reduce the fluorescence. Our circuit will be inserted into E. Coli. We had the idea of placing the bacteria in a gellan bead, as did iGEM team Bristol 2010 , this is a way to enclose our bacteria and to maintain it safe from the environment and another things that could represent a risk for them, as well as to help in the bio safety area because the bacteria would not be in direct contact with the crops; besides it helps the life length of bacteria.

In summary: Vip3Ca3 production is regulated by temperature: it is produced below the 32ºC and at higher temperatures it is turned off.
For more details you can see the circuit: Main Idea Description of the project.