Multicolor detection of combed DNA molecules using quantum dots.
DNA combing is a useful strategy for manipulating single DNA molecules and has a wide range of applications in genetics, single molecule studies, and nanobiotechnology. Visualization of combed DNA molecules is usually performed by using DNA binding organic dyes. Such dyes are not suitable in all circumstances, especially because of their photoreactivity. We have developed a method for the detection of combed DNA molecules by fluorescence microscopy that avoids the use of DNA-staining agents and does not perturb the structure of the DNA molecule. Biotin- and/or digoxigenin-modified DNA fragments are covalently linked at both ends of a DNA molecule via sequence-specific hybridization and subsequent ligation. After the modified DNA molecules have been combed on a polystyrene-coated surface, their ends are visualized by multicolor fluorescence microscopy using conjugated quantum dots.
Escudé
Christophe
C
Régulations et Dynamique des Génomes, INSERM U565, Muséum National d'Histoire Naturelle, Paris Cedex 05, France. escude@mnhn.fr
Géron-Landre
Bénédicte
B
Crut
Aurélien
A
Desbiolles
Pierre
P
D001583:Benzoxazoles; D001710:Biotin; D004247:DNA / Q000032:analysis* / Q000302:isolation & purification; D004076:Digoxigenin; D005456:Fluorescent Dyes; D008856:Microscopy, Fluorescence; D045663:Quantum Dots; D011806:Quinolinium Compounds
D001583:Benzoxazoles; D005456:Fluorescent Dyes; D011806:Quinolinium Compounds; C075296:1,1'-((4,4,7,7-tetramethyl)-4,7-diazaundecamethylene)bis-4-(3-methyl-2,3-dihydro(benzo-1,3-oxazole)-2-methylidene)quinolinium; D001710:Biotin; D004247:DNA; D004076:Digoxigenin