Team:Caltech/Notebook/material/Biofuel Protocols

From 2012.igem.org

(Difference between revisions)

Revision as of 23:16, 17 July 2012

Biofuel Protocols

Ethanol Assay

Ethanol Color Assay Reaction

Materials

water
1.0M bicine (pH 8.0)
40 mM EDTA
10 mM NAD
4.2 mM thiazolyl blue
16.6 mM phenazine ethosulfate
1 mg/mL alcohol dehydrogenase enzyme
ethanol

Method

Make a master mix consisting of:

3.6 mL water
1.2 mL bicine
1.2 mL EDTA
1.2 mL NAD
1.2 ml thiazolyl blue
2.4 mL phenazine ethosulfate

Before running the experiment, add 1.2 mL of alcohol dehydrogenase enzyme.

Pipette 5 uL of sample into the 96-well microtiter plate. Add 95 uL of master mix. Measure absorbance at 570 nm. We found that the maximum absorbance of each well occurred at a different time relative to the concentration, so it is best to run a series of measurements over a period of approximately 4 hours.

Back to Materials and Methods

MOPS Buffer

We used the recipe from Teknova listed here but substituted 10 ml of 20% glucose for 8 ml of 50% glucose. Mix all the following ingredients together to get MOPS buffer:

100 ml 10X MOPS Mixture
10 ml 0.132 M K2HP04
100 ml 10X ACGU
200 ml 5X Supplement EZ
582 ml Sterile H20*
8 ml 20% Glucose

@@ Line 56: / Line 56: @@
      <a style="float:right;" href="https://2012.igem.org/Team:Caltech/Notebook/material#Materials_and_Methods";>Back to Materials and Methods</a>
+<h3><a name="MOPS"; style="color: #000000";>MOPS Buffer</a></h3>
+<p>
+We used the recipe from Teknova listed <a href="http://www.teknova.com/EZ-RICH-DEFINED-MEDIUM-KIT-p/m2105.htm"> here </a> but substituted 10 ml of 20% glucose for 8 ml of 50% glucose.  Mix all the following ingredients together to get MOPS buffer:
+</p>
+<ul>
+<li> 100 ml 10X MOPS Mixture </li>
+<li> 10 ml 0.132 M K2HP04 </li>
+<li> 100 ml 10X ACGU </li>
+<li> 200 ml 5X Supplement EZ </li>
+<li> 582 ml Sterile H20* </li>
+<li> 8 ml 20% Glucose </li>
+</ul>
 </body>