MilliporeSigma
  • Home
  • Search Results
  • Design and characterization of a dual-mode promoter with activation and repression capability for tuning gene expression in yeast.

Design and characterization of a dual-mode promoter with activation and repression capability for tuning gene expression in yeast.

Nucleic acids research (2014-07-25)
Mostafizur Mazumder, David R McMillen
ABSTRACT

Modularity in controlling gene expression artificially is becoming an essential aspect of synthetic biology. Artificial transcriptional control of gene expression is one of the most well-developed methods for the design of novel synthetic regulatory networks. Such networks are intended to help understand natural cellular phenomena and to enable new biotechnological applications. Promoter sequence manipulation with cis-regulatory elements is a key approach to control gene expression transcriptionally. Here, we have designed a promoter that can be both activated and repressed, as a contribution to the library of synthetic biological 'parts'. Starting with the minimal cytochrome C (minCYC) promoter in yeast, we incorporated five steroid hormone responsive elements (SHREs) and one lac operator site, respectively, upstream and downstream of the TATA box. This allows activation through the testosterone-responsive androgen receptor, and repression through the LacI repressor. Exposure to varying concentrations of testosterone (to vary activation) and IPTG (to vary repression) demonstrated the ability to tune the promoter's output curve over a wide range. By integrating activating and repressing signals, the promoter permits a useful form of signal integration, and we are optimistic that it will serve as a component in future regulatory networks, including feedback controllers.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Testosterone, ≥98%
Sigma-Aldrich
D-(+)-Galactose, ≥99%
Sigma-Aldrich
DL-Tryptophan, ≥99% (HPLC)
Sigma-Aldrich
DL-Tryptophan, ≥99% (HPLC)
Sigma-Aldrich
Ammonium-14N2 sulfate solution, 40 wt. % in H2O, 99.99 atom % 14N
Sigma-Aldrich
Adenine, ≥99%
Sigma-Aldrich
Adenine, BioReagent, suitable for plant cell culture, ≥99%
Sigma-Aldrich
Adenine, BioReagent, suitable for cell culture
Sigma-Aldrich
D-(+)-Galactose, powder, anhydrous, BioReagent, suitable for cell culture, suitable for insect cell culture
Sigma-Aldrich
D-(+)-Galactose, BioXtra, pH 5.0-7.0 (20 °C, 1 M in H2O), ≥99%
Sigma-Aldrich
D-(+)-Galactose, ≥98%
Millipore
D-(+)-Galactose, suitable for microbiology, ≥99.0%
Sigma-Aldrich
Ammonium sulfate-14N2 solution, 40 wt. % in H2O, 99.99 atom % 14N
Supelco
Testosterone, VETRANAL®, analytical standard
Sigma-Aldrich
Testosterone, purum, ≥99.0% (HPLC)
Sigma-Aldrich
Ammonium-14N2,sulfate-16O4, 99.99 atom % 14N, 99.99 atom % 16O
Testosterone, European Pharmacopoeia (EP) Reference Standard
Supelco
Galactose, Pharmaceutical Secondary Standard; Certified Reference Material
Supelco
Testosterone solution, 1.0 mg/mL in acetonitrile, ampule of 1 mL, certified reference material, Cerilliant®
Galactose, European Pharmacopoeia (EP) Reference Standard
Adenine, European Pharmacopoeia (EP) Reference Standard
Supelco
Adenine, Pharmaceutical Secondary Standard; Certified Reference Material
Leucine, European Pharmacopoeia (EP) Reference Standard
Tryptophan, European Pharmacopoeia (EP) Reference Standard
SAFC
Galactose, plant-derived
USP
Galactose, United States Pharmacopeia (USP) Reference Standard
Sigma-Aldrich
DL-Leucine, ≥99% (HPLC)
Sigma-Aldrich
Ammonium sulfate, BioXtra, ≥99.0%
Sigma-Aldrich
Ammonium sulfate, for molecular biology, ≥99.0%
Sigma-Aldrich
Nitrogen, ≥99.999%