Manipulation of Corynebacterium glutamicum by gene disruption and replacement

We have developed a system for the genetic manipulation of the amino acid-producing Corynebacterium glutamicum. Gene disruption and replacement were achieved by introducing, via conjugation, Escherichia coli vector plasmids carrying manipulated C. glutamicum DNA fragments. We obtained stable mutants in which the chromosomal lysA gene, encoding meso-diaminopimelate decarboxylase, was interrupted by a chloramphenicol resistance cartridge, or in which an essential internal part of the lysA gene was deleted. The deletion mutants retain neither antibiotic resistance markers nor vector sequences. This strategy is generally applicable to the construction of industrial strains to be used in fermentation processes.