Cytochrome Mutants of Bradyrhizobium Induced by Transposon Tn5

Abstract

Transposon Tn5 was used to mutate Bradyrhizobium japonicum USDA 61N. From over 5000 clones containing Tn5, 12 were selected and purified using a chemical reaction to identify oxidase-deficient clones. Four classes of mutants were identified based on the alterations in cytochromes. Most of the mutants had alterations in more than one cytochrome. Southern hybridization analysis of restricted genomic DNA of a representative strain of each class demonstrated that each mutant had a single Tn5 insert. Thus a single Tn5 insert produced pleiotropic effects on cytochromes. One class, which was totally deficient in cytochromes aa(3) and c, produced ineffective nodules on soybeans. Most of the strains representing the other classes produced effective nodules but exceptions were observed in each class. Bacteroids of the wild-type strain contained cytochrome aa(3). Bacteroids from one class of mutants were totally devoid of cytochrome aa(3). Several of these strains produced effective symbioses indicating that cytochrome aa(3) is not required for an effective symbiosis in this DNA homology group II strain which normally has this terminal oxidase in bacteroids.