The primary structure and base specificity of chicken liver RNase CL1 which has been reported by Miura et al. [Chem. Pharm. Bull., 32, 4053-4060 (1984)] as poly U-preferential RNase, were extensively studied. The sequence study of this enzyme and comparison of the amino acid sequence of the enzyme with homologous RNases from oyster and Drosophila melanogaster suggested that RNase CL1 consists of three peptides with 17, 19, and 163 amino acid residues. The amino acid sequence of these three peptides were identified. The two small peptides are joined to the large peptide by disulfide bridges. The amino acid sequence of RNase CL1 had 62 (31.2%) and 63 residues (31.6%) identical with oyster RNase and D. melanogaster RNase, respectively, and belongs to the RNase T2 family RNase. Reassessment of the base specificity of RNase CL1 found that it is guanylic acid, then uridylic acid-preferential, and not poly U preferential.