Mouse tenascin (TN)-encoding cDNA clones were isolated from a cDNA library of the 2H6GR mammary tumor cell line. Nucleotide (nt) and deduced amino acid (aa) sequences revealed the characteristic primary structure, which begins with a signal peptide and TN unique sequences, follows with 14 1/2 epidermal growth factor (EGF)-like repeats and 13 fibronectin type-III repeats (FN repeat), and concludes with fibrinogen-homologous sequences. Similar to chicken and human TN, the mouse TN cDNA contains five consecutive insertional FN repeats, as well as eight constitutive FN repeats. Three different cDNA clones that may have been generated by alternative splicing of these insertional FN repeats were identified and characterized. Based upon the deduced as sequence, a polyclonal antibody was produced against a synthetic TN peptide. It specifically recognized two TN isoforms of 230 kDNA and 190 kDa in protein extracts of mouse tissues. The tissue distributions of mouse TN mRNAs, revealed by Northern blot analysis, suggest that there is tissue-specific expression of TN isoforms. Two distinct mRNA transcripts (7 kb and 5.5 kg) were detected in brain, skeletal muscle, digestive tract and bladder, but only one was observed in lung, kidney (7 kg) and thymus (5.5 kg). TN mRNA expression was down-regulated 1 month after birth in most tissues. However, the 5.5-kb transcript persisted in cerebellum, thymus, and colon. The spatial and temporal patterns of TN expression seem to be controlled at the level of transcription, because analysis of various tissues by Western blots showed the same pattern as that seen in Northern blots.