Team:Lethbridge/Notebook/Lab Work/September

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(Difference between revisions)
(September 23, 2010)
(September 23, 2010)
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<b>Objective:</b> Characterized catechol by xylE enzyme<br>
<b>Objective:</b> Characterized catechol by xylE enzyme<br>
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<b>Method:</b> Measured absorbance of catechol (275nm) and 2-hydroxymuconate semialdehyde (380nm).
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<b>Method:</b> Measured absorbance of catechol (275nm) and 2-hydroxymuconate semialdehyde (380nm).<br>
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*<b>Protocol:</b>
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*1) Grow cells in M9 minimal medium
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*2) Take 1/10 dilution of cells
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<br>
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*3) Introduce 1&micro;L of 0.05M catechol solution into the cell dilution. (Final concentration of 50&microM;).
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*4) Quench the reaction with 5A% w/v trichloroacetate at certain time points. (0,15sec, 30sec, 45sec, 60sec, 2min).
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*5) Spin down cells.
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*6) Measure absorbance of supernatant.

Revision as of 03:32, 24 September 2010




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September 23, 2010

( JV)

Objective: Characterized catechol by xylE enzyme

Method: Measured absorbance of catechol (275nm) and 2-hydroxymuconate semialdehyde (380nm).

  • Protocol:
  • 1) Grow cells in M9 minimal medium
  • 2) Take 1/10 dilution of cells
  • 3) Introduce 1µL of 0.05M catechol solution into the cell dilution. (Final concentration of 50&microM;).
  • 4) Quench the reaction with 5A% w/v trichloroacetate at certain time points. (0,15sec, 30sec, 45sec, 60sec, 2min).
  • 5) Spin down cells.
  • 6) Measure absorbance of supernatant.