characterization of the genetic element coding for lactose metabolism in Lactococcus lactis subsp. lactis KP3

The Lactococcus lactis subsp. lactis KP3 Lac genetic element was investigated. KP3 is a lactosepositive (Lac +) transconjugant which contains no detectable plasmid DNA. The KP3 Lac genetic element was self-transmissible (Tra +) and encoded a reduced bacteriophage sensitivity (Rbs +) phenotype. Matings of KP3 with a recombination-deficient (Rec −) recipient resulted in Lac + transconjugants which were phenotypically indistinguishable from KP3 and contained a 96-MDa plasmid (pJS96). Phenotypic and physical analyses of pJS96 indicated that it was a deletion derivative of a putative pKB32::pJS88 Lac + Tra + cointegrate. pKB32 is the Lac plasmid and pJS88 is the Tra + Rbs + plasmid in L. lactis subsp. lactis 11007, the donor used in obtaining KP3. The results presented suggest that pJS96 is an episome, since it appeared to replicate both as a plasmid and as an integrated part of the chromosome. Conjugal transfer of chromosomal DNA mediated by pJS96 was not observed. Conjugal transfer of pJS96 resulted in Lac + transconjugants containing plasmids ranging in size from 21 to 90 MDa. Only in Rec + recipients were transconjugants isolated which appeared to contain pJS96 integrated into the host chromosome. Restriction analysis of several plasmids in the 21 to 90 MDa range suggested the deletions were due to intramolecular transposition of a transposable element on pJS96. This report suggests that a selftransmissible episome exists in KP3 and provides an explanation of how plasmids which vary in size yet encode similar phenotypes may be formed and disseminated.