Genomic locus showing integration of pRED1 and pINZ1 into the genome.

<p><b>A</b>) pRED plasmid introduces λ Red proteins (Gam, Exo, and Beta) at <i>mutS</i> locus genome, generating EcSIM strains. <b>B</b>) Heterologous genes can be inserted into the MG1655 genome at the <i>lacZ</i> locus through the pINZ plasmid system, generating EcINZ strains. <b>C</b>) PCR confirmation of successful inactivation of <i>mutS</i> locus by the pRED plasmid system. MG1655 colonies were analyzed for integration of pRED-1 using the following primers: mutSF and Cm_R. Lane M; DNA ladder, lane 1; <i>E. coli</i> MG1655 wild type, lane 2; pRED-2 integrated MG1655. <b>D</b>) PCR confirmation of successful integration of the <i>lacZ</i> locus using the pINZ plasmid system. MG1655 colonies analyzed for integration of pINZ by PCR with primers: Ptac-lacZ and P4R. Lane M; DNA ladder, lane 1; <i>E. coli</i> MG1655 wild type, lane 2; pINZ-2-integrated MG1655.