Team:ETH Zurich/expresults/integrases

From 2014.igem.org

Integrases

The design of our XOR gates was based on integrase logic (ref bonnet). This means, depending on the input molecules integrases can be expressed, subsequently switch a terminator sequence previously blocking gene expression, and then the output gene can be transcribed. The details of this approach are explained here (ref recombinase gates).

In order to characterize the integrase system described above, we first combined the pLux promoter with one of our integrase genes followed directly by mCherry to have a direct measure of expression. Also, this system includes an XOR buffer gate per default blocking transcription of sfGFP. Upon integrase activity and switching the gate into ON-state, the terminator should have been removed and sfGFP should have been expressed. We designed three different constructs for characterization of the recombinases and their cross-activity. However, the measurement of fluorescent proteins, with both a plate reader and a FACS (ref), did not indicate GFP expression even though mCherry was clearly detectable upon induction in platereader experiments.

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