Marker-free Chromosomal Integration of the Manganese Superoxide Dismutase Gene (sodA) From Streptococcus Thermophilus Into Lactobacillus Gasseri

FEMS Microbiol Lett. 2005 May 1;246(1):91-101. doi: 10.1016/j.femsle.2005.03.044.

Abstract

A strategy for functional gene replacement in the chromosome of Lactobacillus gasseri is described. The phospho-beta-galactosidase II gene (lacII) was functionally replaced by the manganese superoxide dismutase (MnSOD) gene (sodA) from Streptococcus thermophilus, by adapting the insertional inactivation method described for lactobacilli [Russell, W.M. and Klaenhammer, T.R. 2001 Efficient system for directed integration into the Lactobacillus acidophilus and Lactobacillus gasseri chromosomes via homologous recombination. Appl. Environ. Microbiol. 67, 4361-4364]. L. gasseri carrying the heterologous sodA gene grew on lactose as efficiently as the wild-type parent. An active MnSOD was expressed in the transgenic strain, and the enzyme migrated on PAGE-SOD activity gels to the same position as that of MnSOD from S. thermophilus. The expression of MnSOD from a single copy of sodA integrated in the chromosome of L. gasseri provided enhanced tolerance to hydrogen peroxide, and extended the viability of carbon/energy starved cultures stored at 25 degrees C. This is the first report showing the successful utilization of the pORI plasmids system to generate marker-free gene integration in L. gasseri strains.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology
  • Bacterial Proteins / analysis
  • Chromosomes, Bacterial / genetics*
  • Gene Expression
  • Hydrogen Peroxide / pharmacology
  • Lactobacillus / enzymology
  • Lactobacillus / genetics*
  • Lactobacillus / growth & development
  • Plasmids / genetics
  • RNA, Bacterial / analysis
  • RNA, Messenger / analysis
  • Streptococcus thermophilus / enzymology
  • Streptococcus thermophilus / genetics*
  • Superoxide Dismutase / genetics*
  • Transformation, Bacterial*

Substances

  • Anti-Bacterial Agents
  • Bacterial Proteins
  • RNA, Bacterial
  • RNA, Messenger
  • Hydrogen Peroxide
  • Superoxide Dismutase