Team:Penn/Communication
PENN iGEM 2015
LIGHT BASED COMMUNICATION
The next step in this three month long process was to finally see if the sender and receiver actually will communicate with each other. Up to this point, all the data collected, especially the approximate calculations made with the conversion sequence, supported the claim that our sender cultures should produce a sufficient amount of light to activate the receiver circuit. Thus with these approximations we set to test our proposed sender-receiver system.
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The figure above shows the arrangement of sender and receiver that the team used to run this experiment. All three strains were tested multiple times with a total of three trials per strain. By measuring the RFP expression, Luminescence expression, and O.D. every 2 hours the following data was procured:
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As can be seen from the data, our approximations and predictions about how the system would function were proven true, the receiver was capable of being induced by bioluminescence from the sender. However, this is not the only outcome worth noting. Looking at the RFP expression induced by each strain separately, it can be noted that SY104 was the strain which resulted in the most RFP over time. This observation falls in line with the team’s prediction before that a sender with sustained expression of light will have a greater and more beneficial effect on a receiver which requires a sustained input, like pDawn.
NOT DONE YET…
Even though, we were able to establish successful communication between sender and receiver in our proposed system, our results still do describe a 100% reliable communication system. We compared our senders approximated intensity output (8uW/cm^2) to what was listed as the saturation intensity of pDawn, around 14uW/cm^2 (Ohlendorf R. et. al. 2010). It is quite clearly that we are not reaching the saturation point with our sender culture yet as result of this comparison. However, we are close, as there is approximately a 3 fold difference between our intensity and the proposed saturated intensity. Thus moving forward it would be imperative to try and improve some parts of our circuits to achieve this 3 fold difference.
In fact, we have previously noticed that the addition of nonanol (a carbohydrate that serves as a substrate for the luciferase reaction) increased the luminescence output by 3 fold. However, it also killed a lot of our bacteria and thus we would not recommend it as a reliable method of solving the issue. However, as we know have become familiar with the workings of pDawn and the three separate strains, we hope to be able to edit these parts (e.g. switch out a promoter, switch the receiver, etc.) to produce a system which contains a sender that will always reliably activate the receiver to saturation.