Allozyme variation was examined in 175 individuals representing 13 putative species of Sargassum from the Great Barrier Reef (GBR), Australia. Allozyme variability was low (five polymorphic loci, 12 alleles total), but differences between most morphological species were observed, and were generally greater than those between different geographical populations of the same taxon. Sargassum decurrens, S. oligocystum, S. parvifolium and S. swartzii ϩ S. aquifolium, were clearly separated by fixed gene differences or major differences in allozyme frequencies, providing sound evidence of these species' reproductive isolation since they were sampled in sympatry. Sargassum siliquosum, S. fissifolium, S. ligulatum, and S. spinifex were not as clearly differentiated, but did form consistent groups, indicating some genetic support for their separation as species. It was clear that S. carpophyllum and S. polycystum are conspecific. Single populations of three different species were thought to be possible misassignments (S. opacum ϭ S. polycystum; S. aquifolium ϭ S. swartzii and S. linearifolium ϭ S. fissifolium), but may be morphological variants of the species with which they clustered. Finally, S. oligocystum (subgenus Eusargassum) clustered with S. decurrens, (subgenus Phyllotrichia) suggesting a revision of subgeneric boundaries in the genus Sargassum is required. Allozyme markers will be useful for species identification of Sargassum within the GBR, as many species are clearly distinguished morphologically for only a short time of the year when they have reproductive organs.