Transformation of Escherichia coli C600 by plasmid DNA pBR322

To investigate the possibilities of Escherichia coli C600-pBR322 plasmid DNA system to be used as host-vector system, the optical conditions for the transformation of Escherichia coli C600 by pBR322 plasmid DNA were examined.Maximum transformation to ampicillin and tetracycline resistance was achieved when cells were harvested from Luria broth at CFU/㎖, followed by washing twice in cold buffer (0.1M NaCl + 5mM + 5mM Tris, pH 7.6.).Optimal conditions for the transformation included a pH of 7.0 and a cell-to-DNA ratio of about CFU/ng of plasmid DNA.The transformation efficiency was highest when cells were exposed to 100 mM in 250 mM KG + 5 mM + 5 mM Tris, pH 7.0, before mixing with DNA. A 60 minutes incubation time for cell + DNA mixtures held on ice produced the maximum number of transformants.At best conditions, transformants were formed.The presence of daltons molecular weight pBR322 plasmid DNA in transformants was confirmed by electrophoresis.