Difference between revisions of "Part:BBa K3463011"
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| − | + | This promoter is activated by binding of BHL via the formation of a BHL/RhlR complex. Before using this sequence in the final system, we wanted to ensure that PrhlR can’t trigger the transcription of the downstream gene without its specific transcriptional activator RhlR. Thus, we cloned this promoter into a plasmid with the reporter gene eGFP (<i>PrhlR</i>-eGFP) and transformed it in the <i>E. coli</i> Nissle strain. We chose this particular <i>E. coli</i> strain because it hasn’t any endogenous production of RhlR. Thus, with or without BHL in the culture media, <i>PrhlR</i> shouldn’t be activated and no eGFP should be produced. | |
| + | So we measured the fluorescence of the bacterial cultures to quantify the expression of GFP as a function of time for two growing conditions : without BHL or with 100µM of BHL. The construct will be validated only if no fluorescence will be detected in both conditions. | ||
Latest revision as of 15:31, 26 October 2020
This promoter is activated by binding of BHL via the formation of a BHL/RhlR complex. Before using this sequence in the final system, we wanted to ensure that PrhlR can’t trigger the transcription of the downstream gene without its specific transcriptional activator RhlR. Thus, we cloned this promoter into a plasmid with the reporter gene eGFP (PrhlR-eGFP) and transformed it in the E. coli Nissle strain. We chose this particular E. coli strain because it hasn’t any endogenous production of RhlR. Thus, with or without BHL in the culture media, PrhlR shouldn’t be activated and no eGFP should be produced. So we measured the fluorescence of the bacterial cultures to quantify the expression of GFP as a function of time for two growing conditions : without BHL or with 100µM of BHL. The construct will be validated only if no fluorescence will be detected in both conditions.