Optimization of a plasmid electroporation protocol for Aeromonas salmonicida subsp. salmonicida
Stéphanie Dallaire-Dufresne1, Jean-Guillaume Emond-Rheault1, Sabrina A Attéré1
1Institut de biologie intégrative et des systèmes, Université Laval, Quebec City, QC G1V 0A6, Canada; Département de biochimie, de microbiologie et de bio-informatique, Faculté des sciences et de génie, Université Laval, Quebec City, QC G1V 0A6, Canada; Centre de recherche de l'Institut universitaire de cardiologie et de pneumologie de Québec, Quebec City, QC G1V 4G5, Canada.
Abstract:
Aeromonas salmonicida subsp. salmonicida is a major fish pathogen. Molecular tools are required to study the virulence and genomic stability of this bacterium. An efficient electroporation-mediated transformation protocol for A. salmonicida subsp. salmonicida would make genetic studies faster and easier. In the present study, we designed the 4.1-kb pSDD1 plasmid as a tool for optimizing an electroporation protocol for A. salmonicida subsp. salmonicida. We systematically tested the electroporation conditions to develop a protocol that generates the maximum number of transformants. Under these optimal conditions (25 kV/cm, 200 Ω, 25 μF), we achieved an electroporation efficiency of up to 1×10(5) CFU/μg DNA. The electroporation protocol was also tested using another plasmid of 10.6-kb and three different strains of A. salmonicida subsp. salmonicida. The strains displayed significant differences in their electro-transformation competencies. Strain 01-B526 was the easiest to electroporate, especially with the pSDD1 plasmid. This plasmid was stably maintained in the 01-B526 transformants, as were the native plasmids, but could be easily cured by removing the selection conditions. This is the first efficient electroporation protocol reported for A. salmonicida subsp. salmonicida, and offers new possibilities for studying this bacterium.
