Description

This composite produces constitutively the CymR regulator protein, that binds the Cumate Operator repressing the transcription from the promoter.

Assembly

The CymR gene was obtained by synthesis and then was assembled with other BioBricks.

Results

CymR expression was confirmed by Western Blot analysis. Our CymR has a SV5 tag at the C-terminus in order to be detected by an anti-SV5 Ab. Its functionality was confirmed through the repression of T5 Cumate Operator-GFP.

The validation of the repression of the protein CymR on the T5 cumate operator was verified through the creation of a plasmid containing a single copy of CymR and the T5 cumate operator upstream the GFP (I13504). We saw that the expression of the GFP was completely repressed in absence of p-cumate but it was active in presence of p-cumate.

Looking forward

The next step will be the integration of two copies of CymR in the bacterial genome in order to regulate the toxin genes carried by the plasmid.

Link to the Registry