Team:Peking S/project/blogic/extension

Boolean Logic


Boolean Logic Synthetic Microbial Consortia| Extension of the Boolean Logic

Extension of the Boolean Logic

It can be anticipated that the absence of feedback mechanisms in Boolean or Non-Boolean gene networks could present a major hurdle for signal correction and modulation in synthetic microbial consortia. Feedback is a mechanism looped back to control a system within itself. Specifically, in systems containing an input and output, feeding back part of the output so as to partially oppose the input is negative feedback, which we make use of in our system. Previous Boolean logic synthetic microbial consortia do not possess any feedback, which limits their potential towards more complex behavior. We have designed a genetic sequential logic circuit with a Boolean-logic based feedback loop as illustrated in Figure 1, which has digitalized performances depending on the previous state of the system.

Figure 1: A genetic sequential logic circuit design with feedback loop. The basic structure is an AND gate, while one input is generated from the output via a NOT gate.

Figure 2: Plasmid design for AND gate with a negative feedback loop. The basic structure of AND gate is on pSB4K5, with plux_inv and pBAD controlling T7ptag and supD, respectively. plux_inv denotes the lux repressible promoter, developed in our quorum sensing inverters. The output portion of our feedback loop locates on pSB1A3, with a T7 promoter leading luxI and gfp, where luxI generates AHL to send back signals as a feedback, and gfp is synthesized to illustrate the strength of output. When the system senses no arabinose, the AND gate is always OFF because of lack of supD, and no GFP would be generated. Nevertheless, an intriguing feature in the feedback circuit is that when our system is induced by arabinose. If previous output is 1, namely gfp is synthesized, so luxI would also be synthesized, subsequently generating AHL among the cells. Then plux_inv would be inhibited by AHL, and the AND gate was switched to OFF state. On the other hand, if previous output is 0, neither gfp nor luxI is synthesized, so that plux_inv is on, and with the presence of arabinose, the output would be shifted to 1 from 0, since AND gate would be ON.

In the previous part of our project, we took a ‘building promoters from ground up’ approach to develop a method to synthesize quorum-sensing repressors. In design, we utilized one of the quorum-sensing repressors to implement feedback signal in order to synchronize the population. Figure 2 illustrates our construction of the plasmids for feedback loop. T7ptag and supD forms the AND gate [1], controlled by plux_inv and pBAD, respectively. Output of our system was illustrated by GFP fluorescence, and the lux system served as feedback pathway.

Function of our feedback loop is presented in Table 2.

Table 2. Sequential logic of AND gate with a negative feedback.

Experimental results

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