Generation of readily transformable Bacillus licheniformis mutants.

A set of mutants was generated by targeted deletion of the hsdR loci of two type I restriction modification systems (RMS) identified in Bacillus licheniformis DSM13. Single as well as double knock-outs resulted in strains being readily transformable with plasmids isolated from Bacilli. Introduction of shuttle plasmids isolated from Escherichia coli was routinely possible when the double mutant B. licheniformis MW3 (DeltahsdR1, DeltahsdR2) was used in transformation experiments. Growth and secretion of extracellular enzymes were not affected in any of the mutants. Thus, along with an optimized transformation protocol, this study makes available an urgently needed transformation system for this industrially exploited species.