From 2010.igem.org
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| - | <h1>Measurement of induction of the GAL1 promoter over time in construct GAL1p-(Npep-GFP)</h1>
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| - | <h3>Aim</h3>
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| - | <p>The aim of this experiment
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| - | To test the response of the GAL1 promoter in the presence of galactose over time, by measuring the expression of GFP, the downstream gene.
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| - | </p>
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| - | <h3>Protocol</h3>
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| - | <p>
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| - | 1. Yeasts transformed with a plasmid carrying the GAL1p-(Npep-GFP) construct was inoculated overnight into 5 ml of synthetic defined (SD) medium with amino acids: his (0.2 %), met (0.2 %), ura (0.2 %), trp (0.2 %) and Raffinose (2 %) as the carbon source. <br><br>
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| - | 2. The following evening 861 µl of this cell culture were sub-cultured into a flask containing pre-warmed SD medium (50 mls) to achieve an optical density at 600nm of 0.3 by 10am the following morning. <br>
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| - | 3. At OD 600 of 0.30, a 1 ml sample was taken to represent the t=0 min sample, and then galactose addded to a final concentration of 0.1 % w/v to begin the promoter induction process. Samples were then taken every 20 minutes thereafter for a period of 170 minutes. All samples were pelleted (13000 rpm, 5min, 4 degrees C), washed once with PBS buffer and stored on ice. Once collected all samples were then dispensed in PBS and diluted by a factor of 1/20 for flow cytometry analysis.
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| - | <br><br>
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| - | <h3>Results</h3>
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| - | <br>
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| - | <p>
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| - | Text
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| - | </p>
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| - | <center>
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| - | https://static.igem.org/mediawiki/2010/0/04/Gal-FACS.jpg
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| - | </center>
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| - | <h3>Conclusion</h3>
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| - | <p>
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| - | Text
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| - | </p>
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| - | </html>
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| - | <br>
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Latest revision as of 14:05, 25 October 2010
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