Helmark, Søren2; Hansen, Michael Edberg2; Jelle, B.3; Sørensen, K.I.3; Jensen, Peter Ruhdal4
1 Center for Microbial Biotechnology, Department of Systems Biology, Technical University of Denmark2 Department of Systems Biology, Technical University of Denmark3 unknown4 National Food Institute, Technical University of Denmark
Plasmid transformation in Leuconostoc carnosum 4010 was analyzed. A successful transformation protocol for L. carnosum was established by modifying an existing protocol for Lactococcus lactis. Several parameters, including the number of generations that the cells had grown at the time of harvest, glycine concentration, the time of incubation for phenotypic expression, and the electrical field strength, were investigated and proved to have influence on the transformation frequency. Electrocompetence was found to be transient and to peak in the early exponential growth phase. Optimized conditions resulted in transformation frequencies of up to 6.7 x 10(5) transformants per microgram of plasmid DNA. A total of five plasmids in L. carnosum were successfully introduced and maintained. Interestingly, we discovered that DNA uptake was of a frequency of 3 x 10(-6) to 19 x 10(-6) transformants per CFU in the absence of an applied electrical field. We concluded that L. carnosum is naturally competent.
Applied and Environmental Microbiology, 2004, Vol 70, Issue 6, p. 3695-3699