Two regions of mouse rDNA were sequenced. One contained the last 323 nucleotides of the external transcribed spacer and the first 595 nucleotides of 18S rRNA; the other spanned the entire internal transcribed spacer and included the 3' end of 18S rRNA, 5.8S rRNA, and the 5' end of 28S rRNA. The mature rRNA sequences are very highly conserved from yeast to mouse (unit evolutionary period, the time required for a 1% divergence of sequence, was 30 x 106 to 100 x 106 years). In 18S rRNA, at least some of the evolutionary expansion and increase in G+C content is due to a progressive accretion of discrete G+C-rich insertions. Spacer sequence comparisons between mouse and rat tRNA reveal much more extensive and frequent insertions and substitutions of G+C-rich segments. As a result, spacers conserve overall G+C richness but not sequence (UEP, 0.3 x 106 years) or specific base-paired stems. Although no stems analogous to those bracketing 16S and 23S rRNA in Escherichia coli pre-rRNA are evident, certain features of the spacer regions flanking eucaryotic mature rRNAs are conserved and could be involved in rRNA processing or ribosome formation. These conserved regions include some short homologous sequence patterns and closely spaced direct repeats.