PENN iGEM 2015
LIGHT BASED COMMUNICATION
Up to this point, all the data collected, especially the approximate luminescence intensity of our sender made with the conversion sequence, suggested the luminescence of sender cultures would be sufficient for successful activation of receiver circuit. The next step was to demonstrate successful sender-receiver communication.
The figure above shows the arrangement of the sender and receiver in this experiment. RFP fluorescence, luminescence, and O.D. at 600 nm was measured every 2 hours. With three trials per strain, the following data was procured:
As suggested in our predictions, the receiver was induced by the bioluminescent signal from the sender. Looking at pDawn activation through RFP expression, SY104 was the strain which resulted in the most receiver activation over time. This observation falls in line with the team’s prediction before that a sender with sustained expression light-production will have a greater effect on a receiver which requires a sustained input, like pDawn, than a sender with a transient luminescence output.
NOT DONE YET…
Even though we demonstrated successful communication between sender and receiver in our proposed system, our results do not show a completely reliable communication system. We compared our senders approximated intensity output (8 uW/cm^2) to the previously recorded intensity for saturation of pDawn (around 14 uW/cm^2) (Ohlendorf R. et. al. 2010). We are not reaching the saturation point with our sender culture yet. However, we are close. There is approximately a three-fold difference between our intensity and the proposed saturated intensity. Moving forward, it would be imperative to improve some parts of our circuits to diminish this three-fold difference.
The addition of nonanal (a carbohydrate that serves as a substrate for the luciferase reaction) was shown to increased the luminescence output by Yagur-Kroll and Belkin (Yagur-Kroll and Belkin, 2010). Though it showed a three-fold difference for certain strains, it reduced OD600 readings. However, as we know uncover more familiar about light-sensitive circuits such as pDawn and the mechanism of luminescent genes in different strains, we hope to tune our system (e.g. swap a promoter, switch the receiver, etc.) to yield a sender that reliably activates the receiver to saturation.