Gamma irradiation of DNA in aqueous solution produces single strand breaks with two types of 3'-termini. These termini have been identified as a simple phosphoryl group and a glycolate moiety attached to the 3'-phosphoryl group via the 2"-OH of glycolic acid. Irradiation of 5'-end labeled poly(dG) produces compounds that comigrate with authentic 3',5'-dGDP and 2'-deoxyguanosine-3'-(phospho-2"-O-glycolic acid-5'-monophosphate by thin layer chromatography and polyacrylamide gel electrophoresis. Irradiation of poly(dGdC) followed by digestion to 3'-nucleotides produces 2'-deoxyguanosine-3'-(phospho-2"-O-glycolic acid). The presence of glycolate termini at some strand breaks may affect intracellular repair of these lesions.