[Reaction of fluorescamine with nitrogen bases in nucleotides and single-stranded synthetic and native...

Reactions of fluorescamine with nucleoside bases have been studied. Fluorescamine reacts with NH2 groups of dA, dG, dC, and the characteristic pyrrolinone spectrum with maximum at 490 nm appears. Fluorescamine does not react with dU and dT. The reaction is pH-dependent, but not linked to protonization of amino groups. pK values for dA, dG, dC are respectively 7.3, 8.4, 8.7. At pH greater than 10 or lower than 3 the fluorescence decreases due to formation of non-fluorescent fluorescamine derivates. The average fluorescence yield of pyrrolinone (the product of reaction of fluorescamine with denatured DNA and single-stranded polymers containing dA) is 2.7-3.5 fold higher as compared to mixture of individual nucleotides. The studied reaction can be used sequencing of nucleic acids.