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The recombination of gene coding for LacI protein prevents the leakage expression of the promoter by inhibiting the Lac promoter completely. Without the input signal (IPTG), LuxI protein and cI protein are not produced, so although the luxR gene under the control of cI promoter transcripts continuously, there is no detectable expression of Green Fluorescence Protein. Under this condition, the expression of LacI and cI protein is under the same logic control, whereas the expressions of LuxR and LuxI protein are controlled by an opposite logic. It is based on the time delay during the transfer between the positive and negative logic control, a biological Flip-Flop is successfully constructed.
The inhibition of Lac promoter is released by the addition of IPTG which results in the simultaneous expressions of LuxI and cI protein. The overexpression of cI protein represses the activity of cI promoter and shut down the expression of LuxR protein. However, the originally accumulated LuxR protein could not be degraded timely and binds to the AHL produced by the LuxI protein, thus initiate the expression of Green Fluorescence Protein.
After a period of degradation, the LuxR protein is consumed gradually and become insufficient to combine with AHL to activate the Lux promoter, thus the Green Fluorescence fade to disappear indicating the output signal has been switch to 0 again.
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