Team:USP-Brazil/InterLab/FlowCitometry
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Flow cytometry analysis
To complement our plate reader assays we perfomed flow citometry analysis of the parts provided for the interlab study. We took 4 different culture time points and analysed it for population fluorescence. And the results obtained were quite interesting!
The expression levels of the Testing Device(TD) 1 were even higher than the ones in the positive control. This effect was clear when using RBS BBa_B0034 , with the mean expression levels showing a profile of "very high", "medium" and "low" activity regarding TDs 1, 2 and 3, respectively. This effect was diminished with the bicistronic element BBa_J364100, with less difference among TDs 4,5 and 6
Curiously, the bicistronic element also reduced the noise of the circuit. It is clear from the comparison of different time points that TDs 4 and 5 show a more homogeneous expression pattern when compared to its "cognates" TD1 and TD2, respectively (which have the RBS R0034). This straightforward analysis highlights the importance of elements beside the promoter, namely RBS, for not only the magnitude of the fluorescence signal, but also for the intrinsic translational noise of the construct. This should be taken into consideration during future part design in order to obtain tightly regulated synthetic circuits.
References and Protocols
