Part:BBa_K2944003

pTDH3-GOx-tPGK1

This coding sequences encodes a constitutive yeast promoter regulating expression of the glucose oxidase enzyme from Aspergillus niger. The gene sequence has been optimized for Saccharomyces cerevisiae. Glucose oxidase catalyzes the oxidation of glucose to D-glucono-1,5-lactone and hydrogen peroxide.

Characterization. Mass Spectrometry.

Activity of Glucose Oxidase was measured reacting the peroxide produce by the catalysis of glucose into gluconate with ammonium molybdate. The product of this reaction produces a yellow color that absorbs light at 405nm. The change in absorbance over time can be measured to determine if glucose oxidase is indeed present in the system.
<img id="team photo" class="logo1" src=" " alt="GOx" width="500">
Figure 1 – Absorbance of Cell Supernatant Over Time in the Presence of 100mM Ammonium Molybdate.

The sample was subjected to a solution of 10mM D-glucose and 100mM ammonium molybdate. The absorbance was measured periodically over a time frame of three hours.

<img id="team photo" class="logo1" src=" " alt="fig2" width="500">
Figure 2 Calibration Curve of the Absorbance at 405nm of Ammonium Molybdate


Table 1- Limit of Detection and Quantification of the Calibration Curve
Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 717
  Illegal BamHI site found at 1027
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1051
  Illegal BsaI.rc site found at 2269