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Convergent Strategies for the Attachment of Fluorescing Reporter Groups to Peptide Nucleic Acids in Solution and on Solid Phase
Author(s) -
Seitz Oliver,
Köhler Olaf
Publication year - 2001
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/1521-3765(20010917)7:18<3911::aid-chem3911>3.0.co;2-1
Subject(s) - peptide nucleic acid , förster resonance energy transfer , molecular beacon , combinatorial chemistry , fluorescence , nucleic acid , oligonucleotide , monomer , dna , peptide , chemistry , solid phase synthesis , biochemistry , polymer , organic chemistry , physics , quantum mechanics
The site‐selective conjugation of peptide nucleic acids (PNA) with fluorescent reporter groups is essential for the construction of hybridisation probes that can report the presence of a particular DNA sequence. This paper describes convergent methods for the solution‐ and solid‐phase synthesis of multiply labelled PNA oligomers. The solid‐phase synthesis of protected PNA enabled the selective attachment of fluorescent labels at the C‐terminal end (3′ in DNA) which demonstrated that further manipulations on protected PNA fragments are feasible. For the conjugation to internal sites, a method is introduced that allows for the on‐resin assembly of modified monomers thereby omitting the need to synthesise an entire monomer in solution. Furthermore, it is shown that the application of a highly orthogonal protecting group strategy in combination with chemoselective conjugation reactions provides access to a rapid and automatable solid‐phase synthesis of dual labelled PNA probes. Real‐time measurements of nucleic acid hybridisation were possible by taking advantage of the fluorescence resonance energy transfer (FRET) between suitably appended fluorophoric groups. Analogously to DNA‐based molecular beacons, the dual labelled PNA probes were only weakly fluorescing in the single‐stranded state. Hybridisation to a complementary oligonucleotide, however, induced a structural reorganisation and conferred a vivid fluorescence enhancement.