Resumen de Detection of Non-B-DNA Secondary Structures by S1 Nuclease Digestion
Marcel.li del Olmo Muñoz, Agustin Aranda Fernández, José Enrique Pérez Ortín, Vicente Tordera
In nature, almost all DNA strands are supercoiled in both prokaryotic and eukaryotic cells. Here, we present two cheap and simple laboratory experiments to analyze the different topological states of DNA and, simultaneously, to detect denatured regions and cruciforms in vitro, using the single-strand specific S1 nuclease. A natural (A+T)-rich region of the 3' region of Saccharomyces cerevisiae FBP1 gene and a DNA (A+T)-rich region in pUC plasmids around the terminator of the ampicillin resistance gene (both capable of undergoing supercoiling-dependent denaturation and therefore sensitive to S1 nuclease) have been used in the experiments. Experimental costs are low, and the small amounts of chemicals and the laboratory equipment used are available in every laboratory.
