Part:BBa_C0060:Experience

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••••• UNIPV-Pavia iGEM 2011

NB: unless differently specified, all tests were performed in E. coli MGZ1 in M9 supplemented medium at 37°C in low copy plasmid pSB4C5. The AiiA enzyme activity has been characterized under the regulation of ptet promoter, assaying its enzymatic activity, with these parts: BBa_K516220 pTet-RBS30-AiiA-TT BBa_K516221 pTet-RBS31-AiiA-TT BBa_K516222 pTet-RBS32-AiiA-TT BBa_K516224 pTet-RBS34-AiiA-TT A system of differential equations has been derived and parameters would have been identified, studying the exponential growth phase: The four measurment parts ptet-RBSx-LuxI were quantitatively characetrized using the BBa_T9002 biosensor. Unfortunately the model parameters (kcat, kM,AiiA) were not estimated because placing the device in low copy plasmid (pSB4C5)resulted in no HSL degradation, as shown in the figures below: A semi-quantitative information about AiiA enzyme activity is given, performing an experiment in E. coli TOP10, with the four measurement devices in pSB1A2; the results are summerized in the graphs below: In this way a measure of AiiA activity is given, as the percentage of HSL degraded in 7 hours (measurements provided as average [stdandard deviation]): Measurement device HSL degraded percentage BBa_K516220 96.11 [3.66] BBa_K516221 77.60 [2.66] BBa_K516222 50.66 [16.87] BBa_K516224 100 [0] In order to evaluate the RBSs efficiency, the ratio between the percentage of the degraded HSL realtive to the measurement device with the standard RBS (BBa_B0034) was computed: RBS RBS efficiency BBa_B0030 0.96 BBa_B0031 0.78 BBa_B0032 0.51 BBa_B0034 1

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