A highly efficient electroporation system for transformation of Yersinia

Add time:08/23/2019    Source:sciencedirect.com

The various pathogenic Yersinia species are not readily and efficiently transformed by classical methods. For this reason, the electroporation technique was applied for genetic transformation of these species. Using optimal conditions, we were able to transform the six Yersinia strains studied with the two most widely used groups of plasmids: pSU2718 (a pACYC184 derivative) and pK19 (a pUC19 derivative). Only Yersinia enterocolitica (Y.e.) serotype 0:8 gave poor results (< 5 × 102 transformants/μg) DNA). Electrical transformation of the other species resulted in high efficiencies, up to 105 transformants/μg DNA for Y.e. serotypes 0:3 and 0:9, 106 for Y. pseudotuberculosis and 107 for Y. pestis. The results varied for each strain with the type of plasmid used. Neither the introduced foreign plasmid nor the resident 72-kb virulence plasmid underwent detectable deletions. Transformation was most efficient with supercoiled DNA, decreasing by one and four orders of magnitude for relaxed circular and linearized plasmids, respectively. The ability to easily and efficiently transfer plasmid DNA via electroporation will greatly facilitate the application of recombinant DNA technology for direct cloning and analysis of significant genes into Yersinia.

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