A Heat Shock Following Electroporation Induces Highly Efficient Transformation of Corynebacterium Glutamicum with Xenogeneic Plasmid DNA

An improved method for the electrotransformation of wild-type Corynebacterium glutamicum (ATCC 13032) is described. The two crucial alterations to previously developed methods are: cultivation of cells used for electrotransformation at 18 degrees C instead of 30 degrees C, and application of a heat shock immediately following electrotransformation. Cells cultivated at sub optimal temperature have a 100-fold improved transformation efficiency (10(8) cfu micrograms-1) for syngeneic DNA (DNA isolated from the same species). A heat shock applied to these cells following electroporation improved the transformation efficiency for xenogeneic DNA (DNA isolated from a different species). In combination, low cultivation temperature and heat shock act synergistically and increased the transformation efficiency by four orders of magnitude to 2.5 x 10(6) cfu micrograms-1 xenogeneic DNA. The method was used to generate gene disruptions in C. glutamicum.