3-step PCR (3S-PCR) protocol for lambda red gene inactivation in Gram-negative bacteria using pKOBEG or other linear DNA recombinogenic plasmids
In collaboration with E. Carniel's group, (Yersinia Unit, Institut Pasteur), we developped a rapid and cloning-free gene disruption method (the 3-Step PCR, see Figure) that allows the extension of the lambda-red strategy to many enterobacteriacea such as commensal and pathogenic Escherichia coli, Yersinia, Serratia, Salmonella, Shigella and Klebsiella. This method, which uses linear DNA fragments where homology regions of 500 bp are flanking a selectable resistance marker, improves the efficiency of the lambda-red method for non E. coli K12 bacteria. A full background and protocole is presented in 4 .pdf documents (see here ). .