Fluorescence intercalator displacement assay for screening G4 ligands towards a variety of G-quadruplex structures |
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Authors: | Tran Phong Lan Thao Largy Eric Hamon Florian Teulade-Fichou Marie-Paule Mergny Jean-Louis |
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Institution: | a University of Bordeaux, ARNA laboratory, F-33000 Bordeaux, France b INSERM, U869, Laboratoire ARNA, IECB, F-33600 Pessac, France c INSERM, U565, Acides nucléiques: dynamique, ciblage et fonctions biologiques, Muséum National d’Histoire Naturelle (MNHN) USM503, CNRS, UMR7196, F-75231 Paris, France d Institut Curie, Section Recherche, CNRS UMR176, Centre Universitaire Paris XI, Bât. 110, F-91405 Orsay, France |
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Abstract: | The potential formation of G-quadruplexes in many regions of the genome makes them an attractive target for drug design. A large number of small molecules synthesized in recent years display an ability to selectively target and stabilize G-quadruplexes. To screen for G4 ligands, we modified a G4-FID (G-quadruplex Fluorescent Intercalator Displacement) assay. This test is based on the displacement of an “on/off” fluorescence probe, Thiazole Orange (TO), from quadruplex or duplex DNA matrices by increasing amounts of a putative ligand. Selectivity measurements can easily be achieved by comparing the ability of the ligand to displace TO from various quadruplex and duplex structures. G4-FID requires neither modified oligonucleotides nor specific equipment and is an isothermal experiment. This test was adapted for high throughput screening onto 96-well plates allowing the comparison of more than twenty different structures. Fifteen different known G4 ligands belonging to different families were tested. Most compounds showed a good G4 vs duplex selectivity but exhibited little, if any, specificity for one quadruplex sequence over the others. The quest for the “perfect” specific G4 ligand is not over yet! |
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Keywords: | Quadruplex DNA-DNA ligand interactions Fluorescence Screening assay |
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