CONDITIONS FOR TRANSFORMATION OF PASTEURELLA-MULTOCIDA BY ELECTROPORATION

Conditions for electroporation of plasmid DNA into Pasteurella multocida were determined for use in developing a cloning system to study virulence factors of P. multocida. The highest efficiency of transformation (1.25 × 107 cfu/μg DNA) was obtained when 7.6 × 1010 cells of P. multocida strain R473 were electroporated at 12.5 kV/cm (10 ms, 5 ng of pVM109). Transformation efficiencies of cells prepared at mid-log-phase were approximately 0.5 log10 lower than early, late, or stationary phases. Neither pBR322 nor pUC-19 were able to transform strain R473 under these conditions, even when DNA concentrations were increased to 1 μg. When pBR322 was ligated with a Pasteurella plasmid, pLAR-1, the hybrid was able to transform strain R473 at an efficiency between 4.5 × 102 and 8 × 104 cfu/μg DNA. Six strains of P. multocida including serotypes A, B, D, and E were transformed successfully. © 1992.