Development of a tightly controlled off switch for Saccharomyces cerevisiae regulated by camphor, a low-cost natural product

Shigehito Ikushima, Yu Zhao, Jef D. Boeke

Research output: Contribution to journalArticle

Abstract

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.

Original languageEnglish (US)
Pages (from-to)1983-1990
Number of pages8
JournalG3: Genes, Genomes, Genetics
Volume5
Issue number10
DOIs
StatePublished - 2015

Keywords

  • CamR
  • Pseudomonas putida
  • Tet system
  • TetR homolog

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Genetics(clinical)

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