Intercalating Nucleic Acids with Pyrene Nucleotide Analogues as Next‐Nearest Neighbors for Excimer Fluorescence Detection of Single‐Point Mutations under Nonstringent Hybridization Conditions

Christensen, Ulf Bech; Pedersen, Erik B.

doi:10.1002/hlca.200390165

Cited by 55 publications

(23 citation statements)

References 28 publications

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“…For example, the introduction of C-nucleotides containing coumarine [39,40] and pyrene [17,18] residues into DNA led to decreases in duplex stability or quenching of fluorescence. Acridine, [41,42] phenanthridinium, [43,44] and pyrene [45][46][47] were incorporated into DNA by means of flexible open-chain 2-deoxyribose analogues. However, fluorescence enhancements in response to matched (or single-mismatched hybridization) were lower than fivefold or have not been reported.…”

Section: Design Principlesmentioning

confidence: 99%

“…However, in previous studies, intercalators have been most commonly linked into DNA helices by means of flexible openchain ribose analogues such as threoninol. [41][42][43][44][45][46][47] Linking base surrogates to flexible open-chain sugar analogues may result in an attenuation of linker effects, which, in contrast, may become apparent when anchoring base surrogates to the rigid 2'-deoxyribose phosphate backbone or its aminoethylglycine equivalent in PNA. Of the six TO base surrogates that were evaluated, TO Q1 proved to be most efficient in stabilizing a PNA-DNA duplex and in conferring selective base pairing of adjacent nucleobases.…”

Section: Duplex Stabilitymentioning

confidence: 99%

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Forced Intercalation Probes (FIT Probes): Thiazole Orange as a Fluorescent Base in Peptide Nucleic Acids for Homogeneous Single‐Nucleotide‐Polymorphism Detection

Köhler¹,

Jarikote²,

Seitz³

2005

ChemBioChem

223

172

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Fluorescent base analogues in DNA are versatile probes of nucleic acid-nucleic acid and nucleic acid-protein interactions. New peptide nucleic acid (PNA) based probes are described in which the intercalator dye thiazole orange (TO) serves as a base surrogate. The investigation of six TO derivatives revealed that the linker length and the conjugation site decided whether a base surrogate conveys sequence-selective DNA binding and whether fluorescence is increased or decreased upon single-mismatched hybridization. One TO derivative conferred universal PNA-DNA base pairing while maintaining duplex stability and hybridization selectivity. TO fluorescence increased up to 26-fold upon hybridization. In contrast to most other probes, in which fluorescence is invariant once hybridization had occurred, the emission of TO-containing PNA probes is attenuated when forced to intercalate next to a mismatched base pair. The specificity of DNA detection is therefore not limited by the selectivity of probe-target binding and a DNA target can be distinguished from its single-base mutant under nonstringent hybridization conditions. This property should be of advantage for real-time quantitative PCR and nucleic acid detection within living cells.

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Section: Design Principlesmentioning

confidence: 99%

Section: Duplex Stabilitymentioning

confidence: 99%

Forced Intercalation Probes (FIT Probes): Thiazole Orange as a Fluorescent Base in Peptide Nucleic Acids for Homogeneous Single‐Nucleotide‐Polymorphism Detection

Köhler¹,

Jarikote²,

Seitz³

2005

ChemBioChem

223

172

View full text Add to dashboard Cite

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“…[2][3][4][5][6][7][8][9] Environmentally sensitive fluorophores can report on binding events such as hybridization and are, thus, principally suited for homogeneous DNA detection. [10][11][12][13][14] Recently, our group and the group of Wagenknecht have introduced intercalator dyes, thiazole orange (TO) [15] and phenanthridinium, [16] respectively, as artificial bases that fluoresce upon hybridization. One of the interesting opportunities provided by such base replacements is to link or even force fluorophores into a specific site of the nucleic acid duplex which under normal circumstances would compete with many other binding sites or even be devoid of binding of fluorophores.…”

Section: Introductionmentioning

confidence: 99%

Divergent and Linear Solid‐Phase Synthesis of PNA Containing Thiazole Orange as Artificial Base

et al. 2005

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Fluorescent nucleobase surrogates provide nucleic acids with interesting properties. We have recently introduced thiazole orange as base surrogate into PNA and found that the so-called FIT (Forced Intercalation of Thiazole orange) PNA probes signal hybridization by enhancements of fluorescence. Common approaches of modifying nucleobases or introducing nucleobase surrogates draw upon the usage of monomer building blocks that have been synthesized in solution phase. The need to prefabricate a base-modified building block can hold up progress if several base modifications or base surrogates are to be evaluated. Herein, a method for divergent solid-phase synthesis is presented that serves the

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“…This strategy has a potential to be used for other fluorescent probes tethering intercalators. [26][27][28][29][30] …”

Section: Introductionmentioning

confidence: 99%

An Efficient Fluorescence Resonance Energy Transfer (FRET) between Pyrene and Perylene Assembled in a DNA Duplex and Its Potential for Discriminating Single‐Base Changes

Kashida

Takatsu

Sekiguchi

et al. 2010

Chemistry A European J

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To increase the apparent Stokes' shift of perylene, pyrene (donor) and perylene (acceptor) were assembled in a DNA duplex to achieve the efficient fluorescence resonance energy transfer (FRET) from pyrene to perylene. Multiple donors were introduced in the vicinity of acceptors through D-threoninol and natural base pairs were inserted between the dyes. Accordingly, donors and acceptors could be accumulated inside the DNA without forming an undesired excimer/exciplex. When two pyrene moieties were located in proximity to one perylene with one base pair inserted between them, efficient FRET occurred within the duplex. Thus, strong emission at 460 nm was observed from perylene when excited at 345 nm at which pyrene has its absorption. The apparent Stokes' shift became as large as 115 nm with a high apparent FRET efficiency (Phi>1). However, the introduction of more than two pyrenes did not enhance the fluorescence intensity of perylene, due to the short Förster radius (R(0)) of the donor pyrene. Next, this FRET system was used to enlarge the Stokes' shift of the DNA probe, which can discriminate a one-base deletion mutant from wild type with a model system by incorporation of multiple donors into DNA. Two perylene moieties were tethered to the DNA on both sides of the intervening base, and two pyrenes were further inserted in the vicinity of the perylenes as an antenna. Hybridization of this FRET probe with a fully matched DNA allowed monomer emission of perylene when the pyrenes were excited. In contrast, excimer emission was generated by hybridization with a one-base deletion mutant. Thus, the apparent Stokes' shift was enhanced without loss of efficiency in the detection of the deletion mutant.

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Intercalating Nucleic Acids with Pyrene Nucleotide Analogues as Next‐Nearest Neighbors for Excimer Fluorescence Detection of Single‐Point Mutations under Nonstringent Hybridization Conditions

Cited by 55 publications

References 28 publications

Forced Intercalation Probes (FIT Probes): Thiazole Orange as a Fluorescent Base in Peptide Nucleic Acids for Homogeneous Single‐Nucleotide‐Polymorphism Detection

Forced Intercalation Probes (FIT Probes): Thiazole Orange as a Fluorescent Base in Peptide Nucleic Acids for Homogeneous Single‐Nucleotide‐Polymorphism Detection

Divergent and Linear Solid‐Phase Synthesis of PNA Containing Thiazole Orange as Artificial Base

An Efficient Fluorescence Resonance Energy Transfer (FRET) between Pyrene and Perylene Assembled in a DNA Duplex and Its Potential for Discriminating Single‐Base Changes

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