As a basis for further comparative studies, nuclear 5S rRNA gene repeats from two plants of the Solanaceae family, tobacco (Nicotiana rustica) and tomato (Lycopersicon esculentum), were isolated and sequenced. The more abundant 5S rRNA gene repeat in tobacco is 430 bp long, while a second less common variant is 521 bp long. In contrast, the 5S rRNA gene repeat from tomato is only 355 bp long. The spacer sequences from these gene repeats, as well as from other published plant nuclear 5S rRNA genes, were compared for repeating or conserved sequence elements. The results indicate that often observed, but non-conserved, repeating sequence elements probably arise spontaneously by unequal crossover with no functional significance. However, three conserved sequence elements immediately upstream of the coding sequence; a C residue at -1, a G + C-rich element centered at -13, and an A + T-rich element centered at -26 resemble regulatory features which have been identified in other types of genes.