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Host-vector system for phenol-degrading Rhodococcus erythropolis based on Corynebacterium plasmids.

M Veselý¹, M Pátek, J Nesvera

¹Institute of Microbiology, Academy of Sciences of the Czech Republic, Vídenská 1083, 14220 Prague 4, Czech Republic.

Applied Microbiology and Biotechnology

|May 24, 2003

Summary

Genetic tools were developed for Rhodococcus erythropolis CCM2595, a phenol-degrading bacterium. A new shuttle vector, pSRK21, enables efficient gene cloning and expression, aiding strain improvement.

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Area of Science:

Microbiology
Molecular Biology
Biotechnology

Background:

Rhodococcus erythropolis CCM2595 is known for its phenol degradation capabilities.
Genetic manipulation is crucial for improving microbial strains for industrial applications.
Existing genetic tools for R. erythropolis were limited, hindering strain development.

Purpose of the Study:

To develop genetic manipulation techniques for Rhodococcus erythropolis CCM2595.
To construct and characterize novel cloning vectors for R. erythropolis.
To enable efficient gene cloning and expression in R. erythropolis for strain improvement.

Main Methods:

Optimization of electrotransformation protocols for R. erythropolis.
Construction of Escherichia coli-Rhodococcus erythropolis shuttle vectors using replicons from Corynebacterium glutamicum.
Characterization of the shuttle vector pSRK21 for cloning sites and stability.
Cloning and expression of the green fluorescent protein (gfpuv) gene.

Main Results:

An efficient electrotransformation procedure yielding up to 7x10(4) transformants/microg DNA was established.
Novel shuttle vectors were constructed, including the small, versatile pSRK21 (5.8 kb).
The pSRK21 vector demonstrated high segregational stability in R. erythropolis and facilitated efficient gfpuv gene expression.

Conclusions:

The developed genetic tools and shuttle vector pSRK21 significantly advance the genetic manipulation of Rhodococcus erythropolis.
These advancements facilitate strain improvement for enhanced phenol degradation and other biotechnological applications.
Efficient cloning and expression capabilities open new avenues for metabolic engineering of R. erythropolis.