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Food-grade cloning and expression system for Lactococcus lactis

C Platteeuw1, I van Alen-Boerrigter, S van Schalkwijk

  • 1Department of Biophysical Chemistry, NIZO, Ede, The Netherlands.

Applied and Environmental Microbiology
|March 1, 1996
PubMed
Summary

New cloning and expression vectors for Lactococcus lactis enable genetic modifications. These vectors utilize a lacF marker for selection and a strong lacA promoter for controlled gene expression.

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Genetic Engineering

Background:

  • Lactococcus lactis is a key bacterium for food fermentation and biotechnology.
  • Efficient genetic manipulation tools are crucial for enhancing L. lactis functionalities.
  • Existing vector systems may have limitations in stability and expression control.

Purpose of the Study:

  • To develop a versatile set of cloning and expression vectors for Lactococcus lactis.
  • To engineer vectors with a strong, controllable promoter and a dominant selection marker.
  • To assess the stability and expression capabilities of the developed vectors.

Main Methods:

  • Construction of expression vectors containing the lacA promoter and pepN terminator.
  • Introduction of the lacF gene as a dominant selection marker in a lacF-deficient L. lactis strain.

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  • Cloning and expression analysis of the Escherichia coli gusA reporter gene under lacA promoter control.

    • Main Results:

    • Developed vectors demonstrated stable maintenance in L. lactis, especially on lactose-containing media.
    • The lacF marker facilitated high-frequency selection of lactose-utilizing transformants.
    • Controlled and high-level expression of the gusA reporter gene was achieved with certain vector constructs.

    Conclusions:

    • The developed vector system provides a robust platform for genetic engineering of Lactococcus lactis.
    • The lacF marker system offers efficient and dominant selection for transformants.
    • These vectors enable controlled gene expression, advancing applications in food biotechnology and synthetic biology.