Gene constructions encoding a leader sequence and the C2, transmembrane, and intracytoplasmic exons of both H-2Ld (pLd-C2) and H-2Dd (pDd-C2) were expressed as cell surface proteins when stably introduced into mouse L cells. Distinct proteins of 17.5 and 16 kd precipitated from pDd-C2 transformants reacted with a C2 domain-specific monoclonal antibody, whereas the pLd-C2 transformants expressed a single 19 kd glycoprotein. A xenoantiserum specific for the exon VI/VIII fusion peptide of H-2Dd precipitated only the 16 kd product, suggesting that this protein was derived from an alternately spliced transcript lacking exon VII of the native gene. CTL specifically recognized the pLd-C2 and pDd-C2 gene products, indicating the existence of CTL reactive against polymorphic determinants in the C2 domain of class I antigens.