IGEM:IMPERIAL/2007/Projects/Biofilm Detector/Design
Infector Detector: Design
Overall Design
Possible Parts
Signaling System
BBa_F1610 RBS-luxI
No promoter - we need to put one on it!
Promoter for Signaling System
BBa_R0040
ptet, if uncontrolled
pLac, if constitutive secretion
pBAD, if controlled secretion
Detection System
BBa_F2620 ptet-LuxR-pLux
Detection range: 1-1000 nM, highest sensitivity from 1-10 nM
Peak secretion: 400 GFP molecules cfu-1 s-1
Peak response time: 27 minutes
Part BBa_T9002 already has GFP attached
GFP Reporter
BBa_EI1354 GFP
Standard GFP, excited/emitted at 501/511 nm, with RBS and stop codons.
LacZ for β-galactosidase assay (yet to be discovered...)
Stage 1: Detector for AHL
Our first aim is to create a system that can accurately detect the concentration of AHL in solution. We found a well characterized detector system sensitive to 3OC6HSL which we will attach to a standard GFP reporter to determine its response to GFP.
We would later test the promoter on a different reporter. The reporter that is chosen for the final comstruct needs to have a long half-life, be stable, be visible to the naked eye and should not have a response time longer than 3 hours.
Stage 2: Detector for bacterial secreted AHL
To more accurately simulate a bacterial biofilm, we decided to create bacteria that would constitutively secrete AHL.
Our detector plasmid would incorporate the Hrp system as an amplifier, to amplify a reporter gene yet to be decided.
