Fig. 1

Construction scheme of the T7 promoter compatible EcN strain. A lacUV5 promoter was added to the 5′ end of the T7 RNA polymerase gene and ligated to a kanamycin resistance cassette flanked by FRT recognition sites. The ligated fragment was amplified via PCR and sequences homologous to the malEFG operon were added to both ends. The fragment containing the T7 RNA polymerase gene, as well as the FRT flanked resistance cassette, was introduced into the chromosome of EcN via homologous recombination using the λ-Red recombinase system. The kanamycin resistance cassette was removed via the FRT recognition sites resulting in the new strain EcN(T7)