FIG. 2.

Strategy for systematic integration of promoters. A master plasmid, pKH966, has two DNA regions for homologous recombination (shaded regions), a kanamycin resistance gene (white arrow), a transcriptional terminator (black arrow), and a cloning site. Each promoter sequence was inserted into EcoRI and XbaI sites of pKH966. A HindIII-BamHI DNA fragment of the resulting plasmid was prepared and introduced into KKS102 by electroporation. The cells are plated onto medium containing kanamycin. Double-crossover allelic exchange is expected to replace the native pE promoter with a constitutive promoter.