DNA sequences of a fragment of nifH from diverse cyanobacteria were amplified, cloned and sequenced to determine the evolutionary relationship of nitrogenase within the cyanobacteria as a group, and to provide a basis for the identification of uncultivated strains of cyanobacteria in the environment. Analysis of 30 nitrogenase DNA and deduced amino acid sequences from cyanobacteria representing five major taxonomic subdivisions showed great variation in phylogenetic distances between the sequences. Sequences from heterocystous cyanobacteria formed a coherent cluster, in which branching forms did not form a clade distinct from the non-branching forms. Nitrogenase sequences from the unicellular cyanobacteria Gloeothece and Synechococcus sp. RF-1 formed a cluster, as did sequences from the genera Xenococcus and Myxosarcina. The nifH sequences of filamentous nonheterocystous cyanobacteria were not closely related to each other, forming deep branches with respect to the heterocystous cyanobacterial nifH sequences. The phylogeny of nifH based on amino acid sequences was consistent with taxonomic relationships among the strains; for example, a sequence obtained form a natural assemblage believed to be dominated by 'Lyngbya' clustered with nifH from Lyngbya lagerheimii. Results also indicate that the phylogeny of nifH among the cyanobacteria is largely consistent with the phylogeny of 16S rRNA, and furthermore that the nifH sequence can be used to identify uncultivated strains of nitrogen-fixing cyanobacteria.