Tissue-specific alternative splicing is an important mechanism for controlling gene expression. Exons 4, 5 and 6 of the human leukocyte common antigen (LCA) gene are included in B cell mRNA but excluded from thymocyte mRNA by differential splicing. In order to study this tissue-specific alternative splicing, we constructed mini-genes that contain only a few of the LCA exons and the SV40 promoter. Mouse B cells and thymocytes were transfected with these mini-gene constructs and the structures of mRNAs were determined by primer extension analysis. The results show that the same primary transcript is spliced alternatively in B cells and thymocytes. This finding suggests that there is a tissue-specific trans-acting factor that regulates the alternative splicing of LCA pre-mRNA. By making various deletion mutants, cis-elements necessary for tissue-specific splicing were confined within the alternatively spliced exons and their immediate flanking intron sequences. Furthermore, linker scanning analysis shows that there are at least three distinct cis-elements within the LCA exon 4 sequence that are required for tissue-specific alternative splicing. Possible mechanisms of LCA alternative splicing are discussed.