Difference between revisions of "Part:BBa K1086002:Experience"
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TMAO enters bacterial cells and generates a cascade of signals, which ultimately activate promoter TorCAD (we have based on Ansaldi et al., 2010 to synthesize TorCAD promoter). We have added mCherry (RFP) downstream of TorCAD in order to measure the promoter’s activation by TMAO. | TMAO enters bacterial cells and generates a cascade of signals, which ultimately activate promoter TorCAD (we have based on Ansaldi et al., 2010 to synthesize TorCAD promoter). We have added mCherry (RFP) downstream of TorCAD in order to measure the promoter’s activation by TMAO. | ||
| − | We made some experiments with E. coli XL1-Blue transformed with this composite to show that our construct works. We added different concentrations of TMAO and measured bacterial cultures fluorescence (excitation: 567 nm; emission: 610 nm) and absorbance (600 nm) for a certain period. The results (Figure 1) show a peak after 3 hours, during exponential phase. | + | We made some experiments with ''E. coli'' XL1-Blue transformed with this composite to show that our construct works. We added different concentrations of TMAO and measured bacterial cultures fluorescence (excitation: 567 nm; emission: 610 nm) and absorbance (600 nm) for a certain period. The results (Figure 1) show a peak after 3 hours, during exponential phase. |
Revision as of 22:44, 26 September 2013
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Applications of BBa_K1086002
We have constructed the composite TorCAD+RFP to detect TMAO (trimethylamine N-oxide) in the serum of patients with cardiac risk. These patients present more TMAO than normal individuals.
TMAO enters bacterial cells and generates a cascade of signals, which ultimately activate promoter TorCAD (we have based on Ansaldi et al., 2010 to synthesize TorCAD promoter). We have added mCherry (RFP) downstream of TorCAD in order to measure the promoter’s activation by TMAO.
We made some experiments with E. coli XL1-Blue transformed with this composite to show that our construct works. We added different concentrations of TMAO and measured bacterial cultures fluorescence (excitation: 567 nm; emission: 610 nm) and absorbance (600 nm) for a certain period. The results (Figure 1) show a peak after 3 hours, during exponential phase.
Conclusion:
Our results show that the composite TorCAD+RFP generates fluorescence in the presence of TMAO.
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