The gene, rpb1, encoding the largest subunit of RNA polymerase II has been cloned from Schizosaccharomyces pombe using the corresponding gene, RPB1, of Saccharomyces cerevisiae as a cross-hybridization probe. We have determined the complete sequence of this gene, and parts of PCR-amplified rpb1 cDNA. The predicted coding sequence, interrupted by six introns, encodes a polypeptide of 1,752 amino acid residues in length with a molecular weight of 194 kilodaltons. This polypeptide contains eight conserved structural domains characteristic of the largest subunit of RNA polymerases from other eukaryotes and, in addition, 29 repetitions of the C-terminal heptapeptide found in all the eukaryotic RNA polymerase II largest subunits so far examined.