Use of stable dye-DNA intercalating complexes to detect cystic fibrosis mutations

Abstract

Several dyes are now known which intercalate into double-stranded DNA with fluorescence enhancement. One such is TOTO (1,1′-(4,4,7,7-tetramethyl-4,7-diazaundecamethylened)-bis-4-[3-methyl-2,3-dihydro-(benzo-1,3-thiazole)-2-methylydidene]-quinolinium tetraiodide), a homodimer of thiazole orange, whose complexes with DNA are stable to electrophoresis and show a 1000-fold enhancement after binding. We demonstrate that this reagent can be successfully used in combination with a laser-excited fluorescence gel scanner to detect the three most common cystic fibrosis mutant alleles, ΔF508, G551D and G542X. This has potential application to mass population heterozygote screening.