PLASMIDS WITH HOMOLOGY TO PFX3 AND PCI3340 IN STARTER LACTOCOCCI AND LACTOBACILLI AND ELECTROPORATION OF SELECTED STRAINS WITH THESE VECTORS

An important consideration in the genetic manipulation of lactic acid bacteria is whether or not strains retain their normal industrial prop erties after transformation with plasmid DNA. A survey of plasmid type s in various lactic acid bacteria showed that pE194-type rolling circl e (RCR) plasmids were present in 24% of 70 commercial Lactococcus lact is cheese starters but not in any of 38 strains representing nine spec ies of Lactobacillus. By contrast, theta plasmids showing homology to the pCI305 rep gene were present in most (though not all) lactococcal strains and not in any of the lactobacilli. Plasmids not belonging to either group were also present in some Lc. lactis and Lactobacillus st rains. Six selected Lc. lactis subsp. cremoris strains were transforma ble with pFX3 (a 4.5 kb pE194-type rolling circle vector), yet four of these strains had significantly lower or zero transformation frequenc ies with pCI3340 (a 5.7 kb pCI305-based theta vector), possibly due to plasmid incompatibility. However, all transformants maintained full m ilk-coagulating activity. In addition to Lc. lactis, strains of Lc. ga rviae, Enterococcus faecalis and Lactobacillus reuteri were able to be transformed with pCI3340, making this a potentially useful vector for these species. (C) 1998 Elsevier Science Ltd. All rights reserved.