Team:Imperial College London/Results

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Revision as of 15:51, 27 October 2010

Department of Bioengineering

Division of Molecular Biosciences

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
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

Experiment 2 \| The threshold value of catechol assay
Aims of experiment \| Results \|

Experiment 3 | Characterizing kinetic parameters of C(2,3)O in whole cells

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
Aims of experiment \| Results \|

Experiment 5 | Characterizing GFP-XylE gene product in whole cell cultures

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

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

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-|style="font-family: helvetica, arial, sans-serif;font-size:2em;color:#ea8828;" align="left"|Experiment 1 | Optimum absorption wavelength for catechol assays
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+|style="font-family: helvetica, arial, sans-serif;font-size:2em;color:#ea8828;" align="left"|                  for catechol assays
 |-
 |valign="top"|'''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