The gene encoding the alpha isoform of rat Ca2+/calmodulin-dependent protein kinase II was cloned, and its exon-intron organization was analyzed. The coding region of cDNA consists of 18 exons spanning more than 50 kilobase pairs. Each of the discrete functional units, such as the ATP-binding site, the autophosphorylation site responsible for Ca2+-independent activity, the calmodulin-binding site, and link structure is encoded by a single exon. The largest and smallest exons consist of 229 and 41 base pairs, respectively. All splice junction sequences flanking the introns conform to the consensus splice junction sequence and the GT-AG splice rule.