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Oligonucleotides containing fluorescent 2'-deoxyisoinosine: solid-phase synthesis and duplex stability

F Seela1, Y Chen

  • 1Laboratorium für Organische und Bioorganische Chemie, Institut für Chemie, Universität Osnabrück, Germany.

Nucleic Acids Research
|July 11, 1995
PubMed
Summary

Incorporating the fluorescent nucleoside 2'-deoxyisoinosine (isoId) into DNA oligonucleotides significantly reduces duplex stability. This fluorescent probe, isoId, shows potential for studying DNA structures and interactions.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Organic Chemistry

Background:

  • The development of modified nucleosides is crucial for advancing nucleic acid research.
  • Fluorescent nucleoside analogs offer unique tools for probing DNA structure and dynamics.

Purpose of the Study:

  • To synthesize and incorporate the fluorescent nucleoside 2 -deoxyisoinosine (isoId) into DNA oligonucleotides.
  • To investigate the impact of isoId incorporation on the thermodynamic stability of DNA duplexes.

Main Methods:

  • Synthesis of phosphonate and phosphoramidite derivatives of isoId.
  • Oligonucleotide synthesis using P(III) solid-phase chemistry.
  • Hybridization of isoId-containing oligomers with complementary DNA strands.
  • Determination of melting temperature (Tm) and thermodynamic data from melting curves.

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Main Results:

  • Incorporation of one isoId residue at the center of a d(T12) oligomer reduced Tm by ~15°C.
  • Incorporation of two isoId residues decreased Tm by ~25°C.
  • The destabilizing effect was largely independent of the opposing natural nucleosides.
  • Structural analysis suggested isoId stacking with dT-dA neighbors and internal loop formation with consecutive isoId residues.

Conclusions:

  • 2 -deoxyisoinosine (isoId) incorporation destabilizes DNA duplexes in a dose-dependent manner.
  • The observed destabilization is primarily influenced by the presence of isoId itself, rather than the complementary base.
  • IsoId's behavior within the duplex depends on its neighboring sequences, potentially forming stacked structures or internal loops.