An origin of transfer from Lactococcus lactis subsp. lactis causes rearrangement in the temperature sensitive plasmids pUCB3522 and pJRS290

Cloning of the oriT region derived from Lactococcus lactis subsp. lactis into plasmids is expected to convert them into mobilizable plasmids in a suitable genetic background. In some cases, however, the cloning of oriT causes deleterious DNA rearrangements. In the work presented, oriT was cloned into two temperature sensitive plasmids, pJRS290 and pUCB3522. The plasmids were transformed into E. coli and electroporated into L. lactis. Restriction digestion of the plasmids derived from the cloned transformed cells showed fragment sizes different from those expected, indicating that rearrangements had occurred. Furthermore, the inability to successfully transform E. coli with one of the plasmids supports this conclusion.