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In Rhizobium trifolii ANU843, host specific nodulation capability is encoded within a 14kb HindIII fragment of the symbiosis plasmid. To gain a better understanding of the regulation of the nodulation (Nod) genes, we have isolated lac operon transcriptional fusions to several genes within this fragment, using the mini-Mu-lac bacteriophage transposon MudI1734. Using a broad-host-range vector, fragments containing MudI1734 insertions were introduced into R. trifolii ANU845, a derivative of ANU843 which lacks the symbiosis plasmid. Four distinct regions were identified within the Nod fragment, insertions in which resulted in nodulation phenotypes similar to those found previously for Tn5. Region I mutants were Nod^- and defective in root hair curling (Hac^-) and corresponded to the nodABC and D genes identified by sequence analysis. Region II mutants showed an exaggerated root hair curling (Hac …