Part:BBa_K3777020
ctcS-tetM-T7(ctcO)-sfGFP
Basic biosensor device for Chlortetracycline detection.
Usage and Biology
The genetic circuit was composed of a coding sequence of Chlortetracycline repressor and tetM genes which was inserted into an expression vectors with a consitive promoter(BBa_J23114) and RBS(BBa_K3777030), as well as sfGFP（BBa_K2762017） under the control of T7 promoter (BBa_K3777006). The terminator we used were BBa_B0010 and BBa_M36305.(Fig 1)
When CTE was absent, TetR would bind to the inducible promoter（PI）and prevent RNA polymerase from initiating transcription, thus repressing the expression of reporter gene. If tet was present, TetR would no longer able to bind to the promoter, resulting in the expression of reporter gene.
We expressed this circuit in the E. coli BL21(DE3) cells for tetracycline detection. Thus we could roughly deduce the concentration of the antibiotics in the sample according to the fluorescence intensity.
Fig.1 Schematic overview of the genetic circuit.
Results
To verify the functionality of the biosensor, we performed a plate-reader experiment and measured optical density and fluorescence intensity every hour. We observed a correlation between concentration of antibiotics in the sample and intensity of fluorescent signal.
The figure shows that our cuircuit can recognize different concentrations of Chlortetracycline, and the fluorescence value was positively correlated with Chlortetracycline concentration which proves that our part can Detect the concentration of Chlortetracycline.
Reference：Alam Khalid K,Tawiah Kwaku D,Lichte Matthew F,Porciani David,Burke Donald H. A Fluorescent Split Aptamer for Visualizing RNA-RNA Assembly In Vivo.[J]. ACS synthetic biology,2017,6(9)
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 3451
Illegal NotI site found at 579 - 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 588
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 2493
Illegal AgeI site found at 2871 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 323
Illegal BsaI.rc site found at 1919
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