The idea of our project is on the basis of studying a whole lot of other projects of former teams. During this process, we find a pretty interesting phenomenon that many projects are involved in 3 major steps of signaling transduction—the sensing, transmitting, and effecting. However, almost all the teams are just constructing a simple signaling transduction system. Some teams did try to construct a more complex system, though, but one single bacterium is not enough to bear the too big plasmid. Unfortunately, their projects therefore failed.
Taking the valuable experience of other teams into consideration, we wonder that maybe we could make it by separating the 3 different functions—sensing, transmitting, and effecting into 3 different bacteria. In this way, the single plasmid is no longer too long and correspondingly the load on our bacteria will be smaller. To this end, we set up the prototype of our project.
In order to construct this kind of system, the key step is arranging the communication of different bacteria. And we name the 3 kinds of bacteria according to their specific function—the sensor, which is used for sensing changes in outside environments; the transmitter, which is focusing on analyzing and transmitting the signals that come from sensor; and the effector, which could execute particular function to the surroundings.
We select several signal pathways to make it possible that the 3 kinds of bacteria communicate with each other. These pathways belong to AHL quorum sensing family (they are Lux, Las, Rhl, Cin pathways, respectively) and the details will be discussed in the following paragraphs.
In the very beginning, we design the gene pathways of the sensor, transmitter, and effector respectively. However, as we investigate further into our project, we find that if we just design a simply single signal pathway within the 3 kinds of bacteria system we are wasting the value of it. Consequently, we change our prime project from the one simply single signal pathway to a more complex double signal pathways.
With the presence of the double signal pathways, here comes some new possibilities. For one thing, you see, one sensor could feel 2 totally different changes in the outside environments and then it will influence the down-stream of our system. Put another way, since there are double signal pathways, we could combine these double signal pathways to construct a complicated logical pathway.
Finally, as we designed originally, we expect that when our sensor receive 2 different changes in the environment or receive these 2 different changes spontaneously our effector could make different actions. These actions are not only limited in the change of level of proteins but also expressing different proteins corresponding to the change of environment.
In the pool of IGEM biobricks we just find 4 available signal pathways (mentioned above, the Lux, Las, Rhl, Cin pathways), whereas inasmuch as our prime design, we need at least 5 signal pathway to finish our gene system. In this situation, we read a whole lot of articles to find a new signal pathway. Fortunately, we find a totally new cell communication signal pathway—the ppy induced Lux-like system.
We manage to get the demanding bacteria—photorhabdus luminescence and we amplify our target gene by PCR. Finally, we add the prefix and suffix of IGEM Competition to form a new biobricks.
Maybe, a little story could help you understand our project clearly.
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