The study of optimal conditions of electroporation in Pseudomonas aeruginosa

A P.aeruginosa strain PA68 isolated from the sputum of a patient suffering from bronchiectasis was used as the recipient strain.Optimum conditions including growth stage of the strain,electroshock voltage,concentration and preservation of competent cell were defined for the electroporation of PA68 with plasmid pSMC28.It was showed that the highest transformation efficiency was up to 1.68×108 CFU/μg DNA under the optimium conditions in which the competent cells were collected at logarithmic growth phase (OD_540=0.7～0.8) and concentrated to about 1011 cells/ml,the mixture of the competent cells and plasmid pSMC28 was eletroporated at 2.6 kV.With this optimal condition,Mu transponson complexes have been successfully transformed into P.aeruginosa strain PA68 and the obtained efficiency was 2.47×104 CFU/μg DNA.This is the first time to electroporate Mu trsnsposon complexes into Pseudomonas spp.The artificial Mu transposons could integrate into bacterial genomes at a single site randomly.Then the phenotype change was the result of the gene inactivation caused by Mu transposon insertion.That will be very helpful for the study of genomic function of Pseudomonas spp.