Transposon Tn5-induced mutagenesis of kanamycin resistance Rhizobium japonicum
Background: Transposase is an enzyme that binds to the ends of a transposon and catalyzes the movement of the transposon to another part of the genome by a cut and paste mechanism or a replicative transposition mechanism. Rhizobium is a genus of Gram-negative soil bacteria, many of which fix nitrogen. Nitrogen fixation is an important part of the nitrogen cycle.
Objectives: The objective of this research was to determine whether at random transposon mutagenesis could be applied in genetic studies of Rhizobium japonicum. One major reason for this is that a range of defined mutations is not available.
Methods: In present study R. japonicum at frequencies sufficient to allow the isolation of large numbers of insertion mutants. The selection of Tn5 mutants was facilitated by the expression, in all the R. japonicum strains we have tested, of the Tn5 encoded kanamycin resistance. A number of auxotrophic and symbiotically defective, single, random transposon (Tn5) mutants were obtained in three slow-growing strains of different DNA homology and serogroups and the single fast growing strain of R. japonicum.
Conclusions: In conclusion, the Tn5 can be used as a generalized mutagen to isolate a variety of mutants with defects in symbiotic nitrogen fixation. The analysis of such mutants should prove to be useful in elucidating the biochemical, genetic, and regulatory events involved in the R. japonicum which effectively nodulates certain Indian soybean cultivars.