Applied Microbiology and Biotechnology

, Volume 65, Issue 1, pp 61–67 | Cite as

A food-grade delivery system for Lactococcus lactis and evaluation of inducible gene expression

  • A. Simões-Barbosa
  • H. Abreu
  • A. Silva Neto
  • A. Gruss
  • P. Langella
Original Paper

Abstract

The genetic improvement of Lactococcus lactis is a matter of biotechnological interest in the food industry and in the pharmaceutical and medical fields. However, to construct a food-grade delivery system, both the presence of antibiotic markers or plasmid sequences should be avoided and the maintenance and expression of the cloned gene should be guaranteed. The objective of this work was to produce crossover mutants of L. lactis with a reporter gene under the control of an inducible promoter in order to evaluate the level of gene expression. We utilized a nuclease gene of Staphylococcus aureus as a reporter gene, PnisA as the nisin-inducible promoter, a non-essential gene involved in histidine biosynthesis of L. lactis as the site for homologous recombination, and pRV300 as a suicide vector for the genomic integration in L. lactis NZ9000. Single- and double-crossover mutants were identified by genotype and phenotype. Relative to episomal transformants of L. lactis, the level of expression of the heterologous protein after nisin induction was similar in the crossover mutants, suggesting that a single copy of the heterologous gene can be used to produce the protein of interest.

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Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  • A. Simões-Barbosa
    • 1
  • H. Abreu
    • 1
  • A. Silva Neto
    • 1
  • A. Gruss
    • 2
  • P. Langella
    • 2
  1. 1.Programa de Pós-graduação em Ciências Genômicas e BiotecnologiaUniversidade Católica de BrasíliaBrasíliaBrazil
  2. 2.Unité de Recherches Laitières et de Génétique AppliquéeInstitut National de la Recherche AgronomiqueJouy en Josas CedexFrance

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