Southern analysis of Volvox genomic DNA revealed two genes homologous to Chlamydomonas reinhardtii alpha-tubulin cDNA. Restriction fragment length polymorphism analysis indicated that the two genes are not genetically linked. Clones representing one of the alpha-tubulin genes have been isolated from a genomic library of Volvox carteri f. nagariensis. A 3153 bp BamHI fragment containing the entire alpha-tubulin gene (1802 bp) plus 707 bp of the 5'- and 644 bp of the 3'-untranslated regions has been sequenced, revealing the following features: (1) the derived alpha-tubulin primary structure of 451 amino acids is highly conserved, differing in two residues from the alpha 1- and in two additional residues from the alpha 2-tubulin of C. reinhardtii; (2) in comparison to the C. reinhardtii genes, the Volvox alpha-tubulin gene contains a third intron; positions of the other two introns are precisely conserved; (3) codon usages are biased towards G or C, and against A, in the third position; 19 codons are absent from the alpha-tubulin coding sequence, and 5 of these are not used in any of 7 compiled Volvox genes; (4) transcription begins with an A, 30 bp downstream of the putative TATA box; upstream of the TATA box is a 14 bp sequence similar to consensus sequences found in all 4 C. reinhardtii tubulin genes and believed to regulate promoter function.