FIG. 4.

catC gene targeted disruption. (A) Plasmid pLK20 was constructed by replacing a central region of 740 bp from the catC gene with the argB gene, used as a selectable marker. The CatC region removed corresponds to amino acids 94 to 341 (Fig. 2). Linear pLK20 was used to transform strain RMS011 to arginine independence. Restriction sites: B, BamHI; E, EcoRI; S, SalI. (B) Total DNA extracted from recipient strain RMS011 and the ΔcatC strain TLK61 was digested with indicated restriction enzymes and used for Southern blot analysis. The probe used was the EcoRI catC fragment from pLK17. The same membrane probed with argB (not shown) gave a hybridization pattern fully consistent with the illustrated integration event. MW, molecular weight (weights are in thousands); WT, wild type.