Difference between revisions of "Part:BBa K1065310"

Figure 1. Different light sources induction power We tested different light sources in order to define the best conditions to switch on and off the circuit. We grew a culture of NEB10beta cells transformed with part BBa_K1065310 until it reached an OD = 0.7 (after, then we split the culture in 4 samples (5ml) and exposed them to different conditions: dark control (the glass tube was wrapped with aluminum foil) (1); blue light bulb (2); white light (3); blue LED light (4);
After an induction period of about 8 hours, in which cultures grew at 37 degrees with stirring, we centrifuged cultures and observed that blue LED, blue bulb and white light all induced successfully the transcription of the reporter, instead the dark control stayed uncoloured. White light worked as an activator, probably because it includes the right wavelength (470 nm). Blue bulb illumination provoked a little less efficient induction.

Figure 2. Our experimental setup involving different types of light

Figure 3 and 4. Enhanced induction upon illumination and prevented transcription in the dark: liquid cultures and pellets. The images show the result of the test after an exposition time of 10 hours. As we can notice from the pellets, blue chromoprotein production occurred only in the blue LED exposed sample (1) and in the one exposed to regular light (2); dark definitely prevented amilCP to be produced (3). However from the liquid cultures we can infer that the blue LED worked more efficiently than the white light.

Figure 5. Absorbance spectra of induced samples and control: Since amilCP is a chromoprotein that absorbes in the UV/VIS range (peak at 588), we pelletted the samples, sonicated and resuspended in 2ml of PBS, then we took some measurements at the UV-VIS spectrometer (PerkinElmer lambda 25) to have some quantitative data of the induction experiment. Data shown are relative to three samples induced for 9 hours. We considered a range between 400 nm and 700 nm. The graph shows that dark (green trace) significantly inhibited the device. It is also confirmed that white light (red trace) somehow induced a little less than the blue LE (blue trace).