We determined the coding nucleotide sequence of phenobarbital-inducible cytochrome P-450 mRNA of the rat by analysis of three cloned cDNAs and by primer extension methods. The deduced amino acid sequence of the cytochrome is composed of 491 amino acids, and its predicted molecular weight and amino acid composition concur with those determined with the purified protein. The sequence of one of the three cloned cDNAs is not identical with that of the other two. In their 922 overlapping nucleotides, 14 nucleotide substitutions occur, and 7 of them result in 6 amino acid changes, therefore indicating the presence of at least two similar but distinct mRNAs for phenobarbital-inducible cytochrome P-450. These substitutions occur in a limited portion of the sequence, apparently forming some sort of a "variable region."