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Team:ETH Zurich/Biology/Validation
From 2011.igem.org
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| - | |style="border-left: none;"|[[# | + | |style="border-left: none;"|[[#AlcR sensor|AlcR sensor]] |
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== Expression of AlcR in ''E. coli'' == | == Expression of AlcR in ''E. coli'' == | ||
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Revision as of 09:21, 21 September 2011
| Validation |
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AlcR sensorExpression of AlcR in E. coliGiven that codon usage is not the same in E. coli and aspergillus nidulans, we analyzed codon usage of AlcR. We obtained a Codon adaptation index (CAI) of 0.7 for the aspergillus nidulans protein in E. coli, especially at the beginning of the gene a few very rare codons were included. To get a fully E. coli-optimized gene, we ordered the gene codon optimized. To make some first test, we exchanged the rare codons at the beginning of the aspergillus nidulans protein with PCR. With these protein we made same first Test of the expression of AlcR. To monitor the expression of AlcR a His-tag was introduced in the test system. western blot Repression test of AlcRTo test our designed AlcR-promotors the following system was designed (see Figure). In present of acetaldeyhde AlcR binds to the AlcR operon and inhibits the expression of GFP. For a better signal assam gfp was used.  results
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XylR sensor |
Xylene degradation |
Bandpass |
References[1] [http://www.ncbi.nlm.nih.gov/pubmed/11550794 Beatrice Felenbok, Michel Flipphi and Igor Nikolaev: Ethanol Catabolism in Aspergillus nidulans: A Model System for Studying Gene Regulation, Progress in Nucleic Acid Research and Molecular Biology, 69: 149-204] |
