Team:Wisconsin-Madison/biosensor

Both the ethanol (EtOH) and n-alkane sensors rely on an arabinose inducible two plasmid system for either EtOH or n-alkane detection. The first plasmid contains genes for producing a protein controlled the pBAD promoter, which turns on transcription of the genes in the presence of arabinose. The second plasmid has the EtOH or n-alkane promoter which turns on the transcription of our RFP. The production of the RFP is then measured with a plate reader.

In our case, it is entirely possible to use a constitutive promoter in place of the current arabinose inducible promoter, as the first part of the system does not rely on the presence of either EtOH or an n-alkane to function. However, by controlling the transcription on the first plasmid via the presence or absence of arabinose, we can a) determine the leakiness of the second plasmid, which relies on the transcription of the first genes, and b) it gives us better control over the activation of our sensor. The pBAD promoter’s activity in the presence of arabinose has a sigmoidal shape; that is, below the saturation concentration of arabinose, the promoter is ‘off’ and above the saturation concentration it is ‘on’ (i.e. there is no gradient dependence on arabinose) It is worth noting, however, that the pBAD promoter is notoriously leaky, and that is may be more effective to suppress its activity with glucose, instead of increasing its activity with arabinose.

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