Replication and maintenance of plasmids in lactococcus lactis

Whereas the majority of plasmids present in lactic acid bacteria, including those of lactococci, are cryptic, several (industrally) important traits are plasmid-encoded and, therefore, can be lost due to plasmid instability. Since plasmid stability is intrinsically linked to the replication of the plasmid, it is clear that a more detailed knowledge of plasmid replication functions is needed for a better understanding of plasmid instability. ... Zie: Summary and general discussion

The Mode of Replication Is a Major Factor in Segregational Plasmid Instability in Lactococcus lactis

The effects of the rolling-circle and theta modes of replication on the maintenance of recombinant plasmids in Lactococcus lactis were studied. Heterologous Escherichia coli or bacteriophage λ DNA fragments of various sizes were inserted into vectors based on either the rolling-circle-type plasmid pWV01 or the theta-type plasmid pAMβ1. All pAMβ1 derivatives were stably maintained. pWV01 derivatives, however, showed size-dependent segregational instability, in particular when large DNA fragments were inserted. All recombinant pWV01 derivatives generated high-molecular-weight plasmid multimers (HMW) in amounts that were positively correlated with plasmid size and inversely correlated with the copy numbers of the monomeric plasmid forms. Formation of HMW or reductions in copy numbers were not observed with pAMβ1 derivatives. The results indicate that HMW formation and/or reduction in plasmid copy numbers is an important factor in the maintenance of pWV01 derivatives. It is concluded that theta-type plasmids are superior to rolling-circle-type plasmids for cloning in lactococci