Journal
G3-GENES GENOMES GENETICS
Volume 5, Issue 10, Pages 1983-1990Publisher
GENETICS SOCIETY AMERICA
DOI: 10.1534/g3.114.012765
Keywords
CamR; Pseudomonas putida; TetR homolog; Tet system
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Funding
- DARPA CLIO program [N66001-12-C-4020]
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Here we describe the engineering of a distant homolog of the Tet repressor, CamR, isolated from Pseudomonas putida, that is regulated by camphor, a very inexpensive small molecule (at micromolar concentrations) for use in Saccharomyces cerevisiae. The repressor was engineered by expression from a constitutive yeast promoter, fusion to a viral activator protein cassette, and codon optimization. A suitable promoter responsive to the CamR fusion protein was engineered by embedding a P. putida operator binding sequence within an upstream activating sequence (UAS)-less CYC1 promoter from S. cerevisiae. The switch, named the Camphor-Off switch, activates expression of a reporter gene in camphor-free media and represses it with micromolar concentrations of camphor.
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