Two cDNAs for possible splicing variants of a 74-kDa regulatory subunit (B" or delta) of human protein phosphatase 2A, were isolated. These variants were identified from human cerebral cortex by library screening and PCR, and designated delta1 and delta3 isoforms, while the previously reported isoform [Tanabe et al. (1996) FEBS Lett. 379, 107-1111 was designated delta2. Compared with the delta2 isoform, the delta1 isoform contained a 32-residue insertion beginning at residue 84, and consisted of 602 amino acids in all. The delta3 isoform lacked a 74-residue sequence corresponding to residues 1083 of the delta2 isoform, and consisted of 496 amino acids. Using isoform-specific antipeptide antisera, the 74-kDa subunit (B" or delta) originally purified from human erythrocytes was identified as the delta1 isoform.