Part:BBa_K2771020
Ratiometric promoter characterization reporter
Combination of EYFP and ECFP to be used in ratiometric characterization of promoters. Both the genes will be exposed to the same external influences (eg. plasmid copy number, culture medium, metabolic burden) and the constitutive expression of ECFP serves as a control for normalization of the EYFP expression, giving the ratiometric output EYFP/ECFP, enhancing the possibility of characterization of intrinsic promoter properties.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
This part is an improvement on a YFP reporter (BBa_E0430) has shown to give better measurements when normalizing YFP output by dividing by CFP fluorescence than when just measuring YFP and dividing that value by the measured OD600. This gives us the possibility of a much more reproducible result, less dependant on growth media and cell density (a rather unreliable parameter to measure) at the time of each measurement. Another good point for using this is that dividing fluorescence measurements gives us an adimensional parameter, that has greater capacity for comparation with other experiments and constructs. In our measurements of quorum-sensing-responsive promoters activity, we compared the variance of our controls, which should have a constant value for fluorescence due to a constant amount of leakiness, when normalizing by the OD600 value and the CFP fluorescence measurement. We found, in this and other experiments, that the normalization using CFP presented significatively less variance. In this example, we found a variance that represented, when reaching equilibrium value, a percentage of the mean twice as small as with the normalization with the OD.
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