Team:UNIPV-Pavia/Methods Materials/Sensing ethanol concentration: our do-it-yourself kit

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• LB medium
• DNA resuspension from iGEM 2009 plates
• Transformation
• X-Gal staining protocol for beta galactosidase (blue/white screening)
• Glycerol stocks
• Plasmid extraction
• BioBrick digestion with restriction enzymes
• Electrophoresis
• DNA gel extraction
• DNA precipitation with sodium acetate
• Ligation
• PCR
• Sensing ethanol concentration: our do-it-yourself kit

In order to sense the presence of ethanol in the culture broth, we initially used a commercial kit, which gave us disappointing results, as not even the calibration resulted linear. Then, inspired by the threat of being caught drunk by the police at the wheel, we looked into whether it was possible to set up a similar testing system to the one used by police officers for breath tests. We discovered that there are two different ethanol sensitive devices: the first type is based on measuring the variation of potential on a charged semiconductor, but we immediately discarded this as it was not possible for us to reproduce a similar device in the short term. The second system looked much more achievable. Actually, it is nothing more than a colorimetric assay, where a redox reaction between a dichromate ion ("http://it.wikipedia.org/wiki/Cromo" \o "Cromo" Cr2 "http://it.wikipedia.org/wiki/Ossigeno" \o "Ossigeno" O72-) and ethanol reduces the chrome’s charge from +6 to +3 and turns the solution from orange to blue in a linear relationship with the amount of ethanol dissolved into the medium.