Genetic transformation of Clostridium botulinum hall a by electroporation

Summary A method was developed for the introduction of plasmids into Clostridium botulinum by electroporation. A 4.4 kb plasmid vector, pGK12, which contains genes for resistance to erythromycin (Em^r) and chloramphenicol (Cm^r) was electroporated into C. botulinum type A (Hall A). The highest transformation efficiency was obtained using midlog phase cells, 10% PEG 8000 as the electroporation solution, and 2.5 kV field strength. The transformation efficiency was highest (10³ transformants/g of DNA) when 1 g of plasmid DNA and 4 × 10⁸ CFU/ml of recipient cells were used. Plasmid DNA recovered from the transformants was indistinguishable from that introduced on the basis of restriction enzyme digestion and agarose gel electrophoresis.