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The nucleotide sequence of a 404-base cDNA encoding the cysteine-rich, tyrosine-containing mouse protamine has been determined. This insert, isolated from a mouse testis cDNA library, encodes a polypeptide of 50 amino acids of which 28 are arginine, 9 are cysteine, and 3 are tyrosine. The insert contains the complete 3' noncoding region of 151 bases and most of the 5' noncoding region. The predicted amino acid sequence of mouse protamine 1 is about 80% homologous to boar protamine and 67% homologous to bull protamine and contains the central, highly basic domain of four arginine clusters found in the trout protamines. The identification of a cDNA clone for a mouse protamine will facilitate studies of the evolution, regulation, and protein-DNA interaction of this nuclear protein unique to haploid spermatogenic cells.