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Related Experiment Videos

Bacteriophage T7 RNA polymerase-controlled specific gene expression in Pseudomonas.

J Davison1, N Chevalier, F Brunel

  • 1Unit of Molecular Biology, International Institute of Cellular and Molecular Pathology, Brussels, Belgium.

Gene
|November 30, 1989
PubMed
Summary

This study extends the T7 phage RNA polymerase system for gene over-expression to Gram-negative bacteria beyond E. coli. Researchers achieved specific protein synthesis in Pseudomonas using compatible plasmids.

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Area of Science:

  • Molecular Biology
  • Microbiology
  • Biotechnology

Background:

  • The T7 phage RNA polymerase system enables exclusive gene over-expression in Escherichia coli.
  • This system relies on genes cloned downstream from the T7 phi 10 promoter.

Purpose of the Study:

  • To demonstrate the applicability of the T7 RNA polymerase system in Gram-negative bacteria other than E. coli.
  • To utilize compatible wide host range plasmids for this extension.

Main Methods:

  • Employing rifampicin (Rif)-resistant T7 RNA polymerase.
  • Using compatible wide host range plasmids for gene cloning and expression.
  • Demonstrating in vivo synthesis and radiolabelling of E. coli galactokinase in Pseudomonas ATCC19151.

Main Results:

Related Experiment Videos

  • Successful extension of the T7 RNA polymerase over-expression system to Pseudomonas ATCC19151.
  • Specific in vivo synthesis and radiolabelling of E. coli galactokinase were achieved.
  • Identified two polypeptides synthesized post-rifampicin treatment in plasmid-free Pseudomonas, indicating stable mRNA species.

Conclusions:

  • The T7 RNA polymerase system is adaptable for gene over-expression in diverse Gram-negative bacteria.
  • This expands the utility of the T7 system for biotechnological applications in various bacterial hosts.
  • The presence of stable mRNA species in Pseudomonas was incidentally observed.