Manipulation and Direct Observation Techniques for Investigation of DNA Metabolic Reactions pp. 373-384
Authors: (Masahiko Oshige, Shinji Katsura, Department of Chemical and Environmental Engineering, Graduated School of Engineering, Gunma University, Gunma, Japan)
Abstract: So far, DNA metabolic reactions such as DNA replication, DNA repair, DNA recombination and transcription, have been investigated intensively; however these studies were carried out by analyzing products of DNA metabolic reactions. Usually, these analyses were carried out by electrophoresis or the incorporation of radioisotope-labeled nucleotides. Because the reactions involve several million molecules, these analyses can only measure average behaviors of a large number of molecules in the reaction, and cannot measure the behavior of individual molecules. Therefore elementary steps of DNA metabolic reactions, such as binding rate, range of continuous DNA synthesis, and the dissociation rate of DNA polymerases still remain unknown. Single molecule observation permits analysis of behavior of individual molecules, and then this method may give new knowledge about DNA metabolic reactions. However, DNA molecules in random-coiled states voluntary shrink due to entropic force, therefore manipulation of DNA configuration is inevitable for measuring DNA length and the mapping of binding sites of factors involved in DNA metabolic reactions. In this article, we mention single molecule observation of DNA metabolic reactions and manipulation techniques for configurations of DNA molecules.
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