Abstract 1. 1. A study has been made of the action of Bacillus subtilis RNAase [polyribonucleotide 2-oligonucleotidotransferase (cyclizing) (B. subtilis)] on a number of polynucleotides, oligonucleotides and TMV-RNA. 2. 2. Poly-I is hydrolyzed by the enzyme more rapidly than Poly-A, which in turn is hydrolyzed more rapidly than Poly-U. Poly-C, by comparison, is almost resistant. In each case the nature of the pattern of the hydrolysis products is much the same. Hydrolysis of the polynucleotide to yield mainly cyclic tri- and cyclic dinucleotides proceeds rapidly. The breakdown of the cyclic trinucleotides is a relatively slow process and the hydrolysis of the cyclic dinucleotide to the cyclic mononucleotide end-product is very slow. In the case of the inosinic acid series there is a measurable hydrolysis of the cyclic mononucleotide to 3′-inosinic acid. 3. 3. B. subtilis RNAase hydrolyzes the dinucleotide GpCp more rapidly than it does triadenylic acid, and the dinucleoside monophosphates GpA, GpC, GpG, GpU, IpC and XpC, more rapidly than diadenylic acid. Dinucleoside monophosphates of the type ApA, CpA, UpA, are resistant to the enzyme. 4. 4. Although Poly-G was not available for these studies it is predicted from the above results that Poly-G would be hydrolyzed by B. subtilis RNAase at a far greater rate than any of the other polynucleotides investigated. 5. 5. The chromatographic pattern of the products resulting from the hydrolysis of TMV-RNA by B. subtilis RNAase is consistent with the pattern expected from a knowledge of the behavior of the enzyme towards the model substrates.