Microtiter-Plate Assay and Related Assays for Nonspecific Endonucleases

Nucleases catalyze the cleavage of phosphodiester bonds in nucleic acids. They range in size from divalent cations, such as Pb2+ , which specifically cleaves tRNAPhe from yeast (1 ), and Mg2+ , which specifically acts on squid tRNALys (2 ) to divalent cations complexed to chelating agents, for example Fe2+ -bleomycin (3 ) and Cu2+ -phenanthroline (4 ) or imidazole (5 ), to complex nucleic acids and proteins, such as ribozymes (6 ,7 ) or the ribonucleoprotein RNaseP (8 ,9 ). Protein nucleases are involved in a variety of important cellular functions, such as DNA restriction, DNA repair and recombination, proofreading of DNA replication, DNA cleavage during programmed cell death and RNA processing, maturation and editing (10 ,11 ). Nucleases may be extremely specific; for example, homing endonucleases, or nonspecific. Examples of the latter include nucleases found in the gastrointestinal tract of higher vertebrates and those secreted into the medium by various microorganisms.