Abstract: A method for characterising nucleic acid molecules comprises the steps of i) introducing a modified base which is a substrate for a DNA endonuclease, for example Endonuclease V from E. coli, into a target DNA molecule; and ii) reacting the nucleic acid containing the modified base with said DNA endonuclease such that the nucleic acid is cleaved to generate an upstream fragment condition said modified base and bearing a 3? hydroxyl group. The upstream fragment generated in step ii) can be used as a primer for a subsequent extension reaction. The modified base as such targets cleavage of the nucleic acid strand because it directs the endonuclease to cut the nucleic acid strand on which it resides at a site predominantly one nucleotide removed therefrom, so that the modified base remains in the nucleci acid as th upstream fragment. The method can be used for detecting polymorphisms and mutations and for nucleic acid indentification and profiling.