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Team:Groningen/PBADmeat
From 2012.igem.org
Our construct idea is simple: production of pigment under the regulation of rotten-meat reactive promoter. When the Bacillus subtilis senses the volatiles from the rotten meat, the rotten meat promoter becomes active thus allowing the production of downstream genes. We put pigment genes available downstream of the promoter so that the pigment would be produced when the promoter is activated.
We use our Bacillus subtilis backbone (BBa_K818000) that has sacA and chloramphenicol resistance gene for chromosomal integration and transformants antibiotic screening. This backbone also has E. coli origin of replication, so it can be amplified inside E. coli.
There are several ideas to improve our current construct, such as: multi-colored pigment system and fine tuning of the pigment production.
The pigment production can be tuned to a desired speed and sensitivity with different regulating promoter, different rbs, and with positive feedback system to increase the pigment production. One example of the positive feedback system that can be applied to increase pigment production under the regulation of the rotten meat promoter:
When the rotten meat promoter is activated, the pigment and inducer will be produced. The positive feedback loop is then formed so that the pigment and the inducer will be in the loop, increasing the production rate of the pigment. This system is meant to increase the production speed of the pigment. One of the possible set of inducible promoter-inducer is pRE promoter (BBa_K116603) with CII (BBa_K116602).
This idea basically mimics the traffic light function: different colors production for every state of the meat. When the meat is still fresh, a specific pigment will be produced. When the meat starts to rot, another pigment will be produced overriding the previous pigment. This can be accomplished by either producing different pigments at different stage of the meat or by producing a specific pigment at fresh meat stage and then produce a compound that reacts to the produced pigment causing it to change color.
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