Plasmid pKD13 was used as the template for amplification of the linear cassette using primers 1 and 2. The kan cassette was removed by expressing the FLP recombinase encoded on plasmid pCP20. Strain NRD204 (K-12 MG1655 araBAD::cat; De Lay and Cronan, 2007) was generously donated by Dr. Cronan (University of Illinois at Urbana-Champaign).