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High-throughput Screening for Chemical Modulators of Post-transcriptionally Regulated Genes
09:44

High-throughput Screening for Chemical Modulators of Post-transcriptionally Regulated Genes

Published on: March 3, 2015

Continuous control of the flow in biochemical pathways through 5' untranslated region sequence modifications in mRNA

Rahmi Lale1, Laila Berg, Friederike Stüttgen

  • 1Department of Biotechnology, Norwegian University of Science and Technology, Sem Sælands vei 6/8, N-7491 Trondheim, Norway.

Applied and Environmental Microbiology
|February 22, 2011
PubMed

Insights

New 5' untranslated region (UTR) variants reduce uninduced expression from the Pm promoter in E. coli. These variants maintain inducibility for applications in synthetic biology and metabolic engineering, enabling precise control over gene expression.

Area of Science:

  • Molecular Biology
  • Synthetic Biology
  • Metabolic Engineering

Background:

  • The inducible Pm promoter offers tunable gene expression but has uninduced background levels.
  • Reducing background expression is crucial for applications like metabolic engineering and synthetic biology.

Purpose of the Study:

  • To develop 5' untranslated region (UTR) variants of the Pm promoter to reduce uninduced expression.
  • To assess the impact of these UTR variants on gene expression and inducibility.

Main Methods:

  • Doped oligonucleotide mutagenesis and selection were used to create UTR variants.
  • Reporter gene assays (β-lactamase, luciferase, phosphoglucomutase) in Escherichia coli were performed.
  • Variants were tested with the Pm promoter and a constitutive P1 promoter.

Main Results:

  • Five UTR variants significantly reduced uninduced expression of reporter genes.
  • The ratio of induced to uninduced expression was maintained or increased.
  • UTR variants showed similar effects on a different promoter (P1), indicating broad applicability.
  • Two variants eliminated uninduced sarcinaxanthin production in engineered E. coli.

Conclusions:

  • UTR engineering is an effective strategy to minimize background gene expression from inducible promoters.
  • These UTR variants enhance control for applications in synthetic and metabolic engineering.
  • The developed UTR variants offer a valuable tool for precise gene expression regulation.

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