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Team:Imperial College London/Results
From 2010.igem.org
| Experimental Results | Exp 1 | Exp 2 | Exp 3 | Exp 4 | Exp 5 | Exp 6 | Exp 7 |
| Testing is a fundamental stage of the engineering design cycle and is a crucial part of charactrising BioBrick Standard Biological Parts so that other people can benefit from our work. We've compiled all our results on this page, detailing how the experiments were carried out and the significance of the data. | |
| Experiment 1 | Optimum absorption wavelength |
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| for catechol assays | |
| Aims of experiment | In this experiment spectra from XylE expressing cells and XylE negative are compared after catechol addition to identify optimum absorption wavelength for quantification of output signal production
Results | Overlay of the spectra from the two cell cultures after catechol addition, reveals that in XylE expressing cultures a broad peak appears with maxima at 380nm | |
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| Experiment 2 | The threshold value of catechol assay |
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| Aims of experiment |
Results | |
| Experiment 3 | Characterizing kinetic parameters of C(2,3)O in whole cells |
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| Aims of experiment | In this experiment we attempt to characterize the kinetic parameters of catechol(2,3) dioxygenase enzyme, an existing registry part in whole cell cultures
Results | Very useful information were acquired out of these assays of which the most important being the catechol(2,3)dioxygenase reaction profile with catechol. The graph generated delineate the course of the reaction in terms of yellow product production over time at various initial catechol concentrations |
| Experiment 4 | Assaying cell-growth in presence of Catechol |
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| Aims of experiment |
Results | |
| Experiment 5 | Characterizing GFP-XylE gene product in whole cell cultures |
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| Aims of experiment | In this experiment we wanted to test if our novel designed biopart, GFP- C(2,3)O fusion protein was functioning under the specification set by the team when designing this inactivated reporter enzyme
Results | Comparing reaction profile graphs of cell cultures of XylE expressing cells Vs GFP-XylE expressing cells showed that the designed reporter enzyme has more than 10 fold decreased activity in comparison to wild type reporter enzyme catechol(2,3)dioxygenase |
| Experiment 6 | In vitro characterization of C(2,3)O in cell lysate |
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| Aims of experiment | In this experiment we attempt to characterize in vitro the kinetic parameters of the enzyme catechol(2,3)dioxygenase in cell lysate
Results | Construnction of Michaelis-Menten curve and determination of kinetic parameters of catechol(2,3)dioxygenase enzyme in E.coli. The calculated Km value, the substrate concentration at which velocity is half of the maximum, is 0.71mM of catechol. The Vmax is 3.37mM/min at x20fold dilution of cell lysate |
