To define the structure of canine B creatine kinase, clones were isolated from a library prepared from dog brain mRNA and constructed in the vector lambda gt11. The entire coding portion, the complete 3' nontranslated region, and 43 bp of the 5' noncoding region are reported. Comparison of the predicted amino acid sequence of canine B creatine kinase with the sequence of canine M creatine kinase shows 81% identity. When compared to cDNAS encoding B creatine kinase isolated form other species unusual and striking nucleotide sequence identity in the 3' noncoding region is present. Moreover, two B creatine kinase clones (BCK2 and BCK38) demonstrate microheterogeneity within the 3' nontranslated region indicating variable processing of B creatine kinase pre-mRNA or the existence of multiple genes encoding canine B creatine kinase.