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Our project aims to control the concentration of living bacteria in a culture. To do so, we designed a genetic network allowing light controlled cell growth. In addition, we built a device in order to send and receive light signals from the bacterial culture. We have thus to create a means of communication from cell to machine and from machine to cell. For cell to machine communication, we chose to measure the red fluorescence of KillerRed. A first function of our device is to excite and measure fluorescence intensity thanks to a light source, excitation/emission optics and a photodiode. In this way, our bacteria will be able to talk to our device. For machine to cell communication, we will use red light to activate light-inducible promoter that triggers KillerRed production and white light to generate ROS thanks to KillerRed phototoxic activity. In our system, the rate of KillerRed production and the number of living cells will be controlled by the intensity of the red and white light beams. A second function of our device is therefore to generate controlled light intensities at different wavelengths. In this way, our device will be able to talk to our bacteria.
Logical scheme of our device The computer is giving orders to the microcontroller that control the light source (intensity and wavelengths) that illuminate the sample. Then the photodiode is measuring the level of fluorescence of the Red Fluorescent protein that is produced in the sample and the microcontroller is giving back the information to the computer that will change the intensity and the wavelength according to the measurement
Our device is built in such a way that the user just needs to input the concentration of living cells he wants and put the Erlenmeyer with our engineered bacteria. From that moment the device works in standalone mode. Firstly our device measures the red fluorescence of the culture that will be the baseline. Then it induces the KillerRed protein using the red light inducible promoter. Every 5minutes, Talk’E.Coli measures the level of red fluorescence of the culture and consequently of KillerRed. With these measurement and the parameters given by the model it can calculate the moment when it needs to activate the ROS emissions with white light to stabilize the living cell concentration. The main asset of the device is that even if there is a lower increase of the concentration of KillerRed or if the killing rate is higher since it measures every 5 minutes the red fluorescence it can adjust exactly the light intensity to correct these rates.
Firstly we will explain the choice of the different components, then the several experiences we did to find the most accurate parameters for each part of the device : the photodiode and Arduino, fluorescence measurement, the electronic circuit, the servomotor. All these elements were then integrated in the box that we designed and built.
