Team:BHSF ND/Demonstrate
2019 IGEM BHSF_NDDemonstrate
Stage 1: Pre-process
Before the engineered bacteria being processed, the microbes are in stage 1. There is no inducer to the transcription factor; therefore, the inducible promoter module is inoperative. To prevent the unintentional leakage, there is constant inhibition caused by the sRNA (the orange sRNA1 and sRNA2) that inhibits the expression of the RBS (the red semicircle) and repressor (the blue R1 and R2). These are the inhibition stages (“0” state) for both of the bistable modules.
Stage 2: Processing
There is an inducer (the yellow ball) to the inducible promoter module that initiates the expression of the following gene. The real input signal starts the expression of the repressor 1 (R1), and the repressor further inhibits the first P1. The expression of the repressor 1(R1) changes the inhibition stage (“0” state) of the first module to the function stage (as 1). The repressor 1 (R1) also inhibits the second P 1, so the second bistable module is still on inhibition (a”0” state)
Meanwhile, the expression of the recombinase in the first bistable module causes the re-direction of the second P1 by the change of attB and attP. The inhibition of the second module prevents the unintentional leakage of the promoter of the sRNA one.
Stage 3: Steal
After the engineered bacteria being stolen, the microbes are in step 3. Bacteria now leave the factory environment after stealing, and the inducer from the factory environment is absent. The lack of inducer to the inducible promoter module causes the weaker expression of the repressor 1 (R1) and the relatively stronger inhibition to R1 caused by the constant sRNA1 expression.
The falling of the repressor 1 (R1) also leads to weaker inhibition to the second P1. The expression of the second P1 as a real, strong signal allows the expression of the following gene. The expression of the repressor 2 (R2) inhibits the expression of the sRNA 2, and the expression of the toxin commits the suicide program. Eventually, the toxin destroys the engineered bacteria, detecting steal and preventing negative effects on factory.
