title: Direct visualization of the quadruplex structures in human chromosome using FRET: application of quadruplex stabilizer and duplex-binding fluorophore.
abstract: The G-quadruplex structures in the telomere of a chromosome can not only protect the internal chromosome sequences by preventing the improper activation of DNA-damage-response pathways but also become targets for cancer treatments. In this manuscript, we wish to prove the existence of G-quadruplex structure formation, rather than G-quadruplex sequence, in chromosome of human cancer cells. Based on our studies, the fluorescent mapping of G-quadruplex structures in the chromosome is possible with the combination of G-quadruplex targeting fluorophore (BMVC, 3, 6-bis-(1-methyl-4-vinylpyridinium)-carbazole diiodide) and duplex-binding fluorophores (Hoechst or propidium iodide). By means of an applicable incubation time between cell cycle period and proper staining procedure to the chromosome, FRET (fluorescence resonance energy transfer) between G-quadruplex targeting fluorophore and duplex-binding fluorophore can increase the signal contrast of the fluorescent color and the fluorescent mapping of quadruplex structures can be easily observed using fluorescence microscopy. These observations are further supported by basic spectral analysis, titration binding assay, gel electrophoresis binding competition assay and confocal microscopy.
authors.affiliation: Graduate Institute of Biomedical Engineering, National Chung Hsing University, 250 Kuo Kuang Road, Taichung 402, Taiwan, ROC.
authors.forename: Yung-Chieh
authors.initials: YC
authors.lastname: Chan
authors.forename: Jyun-Wei
authors.initials: JW
authors.lastname: Chen
authors.forename: Sheng-Yuan
authors.initials: SY
authors.lastname: Su
authors.forename: Cheng-Chung
authors.initials: CC
authors.lastname: Chang
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use; D002227:Carbazoles / Q000737:chemistry; D002877:Chromosomes, Human / Q000648:ultrastructure*; D004249:DNA Damage / Q000235:genetics; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000737:chemistry; D054856:G-Quadruplexes; D006801:Humans; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology; D016615:Telomere / Q000648:ultrastructure
chemical_list: D000970:Antineoplastic Agents; D002227:Carbazoles; D005456:Fluorescent Dyes; C041514:carbazole