HIGHLY EFFICIENT INTEGRATION OF FOREIGN DNA INTO THE GENOME OF THE GREEN SULFUR BACTERIUM, CHLOROBIUM-VIBRIOFORME BY HOMOLOGOUS RECOMBINATION

Highly efficient and reproducible transformation of Chlorobium vibriof orme with plasmid DNA has been achieved by electroporation. Specific p arameters have been optimized for the electrotransformation procedure. The method was developed using a construct containing a full copy of the pscC gene encoding the cytochrome c(551) subunit of the photosynth etic reaction center complex and the aadA gene encoding streptomycin r esistance as selectable marker. Southern blotting analysis showed that the tested colonies were true transformants with the plasmid integrat ed into the genome by single homologous recombination. No transformant s were obtained using the vector without the pscC gene showing that th is vector does not replicate in C. vibrioforme. Thus transformation is possible only by homologous recombination. When using constructs desi gned to inactivate the pscC gene by insertion no transformants were ob tained, indicating that the gene is indispensable for growth. The vect or pVS2 carrying genes for erythromycin and chloramphenicol resistance was shown to replicate in C. vibrioforme. The two transformations sho wn here, provide an important genetical tool in the further analysis o f structure and function of the photosynthetic apparatus in green sulf ur bacteria.