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CREATING THE GENETIC CIRCUIT
From 2012hs.igem.org
Creating the Genetic Circuit
Once we had our idea solidified, the next step was coming up with a circuit to make our idea into a reality. Using the [http://www.tinkercell.com/ TinkerCell] program to create an example of the cell, we wanted to follow the KISS rule and keep things as simple as possible. The 2010 METU iGEM team created a [http://partsregistry.org/wiki/index.php?title=Part:BBa_K352014 part] that successfully detected carbon monoxide, and contained a Lacl promoter, RBS, and CooA, but from that point what the output was seemed to be fairly open for change. We wanted the output smell to be very unpleasant and trigger a negative response, and decided indole, the smell naturally made by e. coli, would be an excellent choice. We chose tnaA, the indole producing gene, as the last major step in our genetic sequence, and an [http://partsregistry.org/wiki/index.php?title=Part:BBa_J45999 odorless chassis] so that indole wouldn't be naturally produced by the detector.
Obstacles in creating the circuit were finding a tnaA producing gene (still working on that) and...
The parts chosen were all marked as 'working' in the parts registry, and most had the same assembly methods, making them compatible. Our major concerns are that we won't be able to find a tnaA gene by itself, or that the parts chosen won't work be compatible in practice.
Also, special thanks go out to our iGEM advisers, Alyssa Henning, !
Parts We are Considering
Chasis: [http://partsregistry.org/wiki/index.php?title=Part:BBa_J45999 BBa_J45999]
CooA: [http://partsregistry.org/wiki/index.php?title=Part:BBa_K352014 BBa_K352014] (CooA coupled with placI and RBS)
tnaA: (for indole production)
- issue: we don't know if we can find a tnaA gene
three genetic circuits we are considering: one with GFP one with indole one with banana.
Protocol
We are considering the following protocols for preparing our competent cells and preforming our transformation.
[http://partsregistry.org/Help:Protocols/Competent_Cells Competent Cells]
[http://partsregistry.org/Help:Protocols/Transformation Transformation]
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