Cloning and Expression in Pseudomonas putida of Two of the Histidine Utilization Genes from Rhizobium fredii

Abstract.

Two of the genes encoding histidine utilization (hut) in Rhizobium fredii strain HH303 have been cloned in Pseudomonas putida and partially characterized. Molecular cloning of the genes was achieved by mobilizing an R. fredii cosmid library into a mutant strain of P. putida containing a Tn5 element in its histidase (hutH) gene. A number of overlapping clones were identified, all of which contain a 7.1-kbp HindIII fragment. The origin of this 7.1-kbp fragment from the chromosome of R. fredii was confirmed by Southern blotting and hybridization studies. In addition, this fragment and the two adjacent HindIII fragments of 9 and 12 kbp respectively were subcloned into pBluescript KS+ and further characterized. Although the R. fredii clones expressed histidase in Pseudomonas, they did not in Escherichia coli, suggesting that E. coli is not a suitable cloning host for Rhizobia genes.