Oligonucleotides functionalized by fluorescein and rhodamine dyes: Michaeladdition of methyl acrylate to 2 '-deoxypseudouridine

The 2'-deoxypseudouridine (5) was functionalized at N(1) with methyl acryla te by Michael addition. The resulting methyl 2'-deoxypseudouridine-1-propan oate (6) was converted to the phosphoramidite 8 and to the amino-functional ized derivative 9, which was transformed into the fluorescein-labeled phosp horamidites 14 and 16. Fluorescent oligonucleotides were synthesized either from these building blocks or by post-synthetic modification of oligomers containing 2'-deoxypseudouridine subunits. The stability of oligonucleotide duplexes was determined from the melting profiles, measured by UV- or VIS- light absorbance, as well as from the fluorescence emission spectra. While small spacer residues did not affect the thermal stability of the 2'-deoxyp seudouridine-containing duplexes, large dye residues led to destabilization .