Perylene Attached to 2′-Amino-LNA: Synthesis, Incorporation into Oligonucleotides, and Remarkable Fluorescence Properties <i>in Vitro</i> and in Cell Culture

Astakhova, Kira; Korshun, Vladimir A.; Jahn, Kasper; Kjems, Jørgen; Wengel, Jesper

doi:10.1021/bc800202v

Cited by 52 publications

(56 citation statements)

References 101 publications

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“…High fluorescence quantum yield, red-shifted emission compared to pyrene (350-500 nm and 420-620 nm for pyrene and perylene in cyclohexane, respectively 23 ), and the remarkable photostability of perylene makes it an attractive label for fluorescence-based detection both in vitro and in vivo. We previously introduced 2′-N-(perylen-3-yl)carbonyl-2′-amino-LNA modification which induced high thermal stability of DNA:DNA and DNA:RNA duplexes, high Watson-Crick mismatch selectivity, red shifted fluorescence emission compared to pyrene, and high fluorescence quantum yields 25 . Additionally, the perylene-LNA probe was successfully applied for detection of mRNA in vivo providing an excitation wavelength which completely eliminated cell autofluorescence 25 .…”

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confidence: 99%

Fluorescent oligonucleotides containing a novel perylene 2′-amino-α-L-LNA monomer: Synthesis and analytical potential

Astakhova

Kumar

Wengel

2011

Collect. Czech. Chem. Commun.

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Dedicated to Professor Antonín Holý on the occasion of his 75th birthday in recognition of his outstanding contributions to the area of nucleic acid chemistry.Herein, a novel fluorescent nucleotide analogue, perylene-2′-amino-α-L-LNA, has been prepared and studied within synthetic oligonucleotides of different sequences. The phosphoramidite reagent was synthesized in 85% overall yield starting from 2′-amino-α-L-LNA nucleoside. Incorporation efficiency of the resulting perylene-2′-amino-α-L-LNA monomer (T*) into synthetic oligonucleotides was significantly improved by replacement of the typically used 1H-tetrazole activator with pyridine hydrochloride. Generally, oligonucleotides containing monomer T* showed high binding affinity towards complementary DNA and RNA targets, batochromically shifted excitation/emission wavelengths with respect to the often applied polyaromatic hydrocarbon pyrene, high fluorescent quantum yields and very low target detection limits (5-10 nM). Fluorescence of single stranded LNA/DNA mixmer oligonucleotide having two incorporations of monomers T* was quenched (quantum yield Φ F = 0.21) relative to duplexes of this probe with complementary DNA and RNA (Φ F = 0.42 and 0.35, respectively). On the contrary, a strong fluorescence quenching upon target binding was demonstrated by two short oligonucleotides of analogues sequences containing monomers T* at 5′-and 3′-terminal positions. We explain the hybridization-induced light-up effect observed for double-labeled probe by a reduction of fluorescence quenching due to precise positioning of the fluorophores within the double-stranded complexes. Furthermore, we propose that a covalent link between two T* monomers in the double-labeled probe provides a remarkable degree of rigidity in the double helix which enforces positioning of the bulky perylene moieties in the nonpolar groove resulting in reduced fluorescence quenching.

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confidence: 99%

Fluorescent oligonucleotides containing a novel perylene 2′-amino-α-L-LNA monomer: Synthesis and analytical potential

Astakhova

Kumar

Wengel

2011

Collect. Czech. Chem. Commun.

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“…Phosphoramidite reagents 5, 41 6 42 and 9 43 were synthesized following published procedures; phosphoramidite reagent 3 was prepared as described below. Pyrene-1-and pyrene-4-carbaldehydes were obtained from commercial suppliers (Sigma-Aldrich, cat.…”

Section: Patients Materials and Methodsmentioning

confidence: 99%

“…Previously investigated (pyrene-1-yl)methyl 2'-amino-LNA (monomer M 1 ) was incorporated into oligonucleotides as a reference for the new FRET donor M 2,41 monomers M 3 and M 4 were prepared as previously described. 42,43 For initial evaluation of pyrene-perylene FRET and SNP diagnostic potential, monomers M 1 -M 4 were incorporated in a model dual-probe FRET system ON1-ON4 and covalently linked double-labeled analogs ON5-ON8 adopted from previous research on PAH-LNA monomers ( Table 1). 29,30 The model FRET probes were prepared using automated phosphoramidite DNA synthesis and first characterized by their melting temperatures with complementary and mismatched DNA/RNA targets (T m ; Tables S1-S6).…”

Section: Synthesis Of Monomer Mmentioning

confidence: 99%

Rapid genotyping using pyrene−perylene locked nucleic acid complexes

Kumar¹,

Myznikova²,

Samokhina³

et al. 2013

Artificial DNA: PNA & XNA

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“…Perylene-LNA/DNA probes have additional applications in the detection of RNA by fluorescence microscopy in cell cultures ( Figure 3A,B) [25]. Short oligonucleotide probes containing perylene-LNA modification 2 and additional oxo-LNAs 1 specifically revealed the presence of target mRNA in fixed cells at excitation/emission wavelengths which completely excluded the cell's autofluorescence (Figure 3).…”

Section: Enzyme-free Hybridization Assays: Specificity In Focusmentioning

confidence: 99%

Novel Signal-Enhancing Approaches for Optical Detection of Nucleic Acids—Going beyond Target Amplification

Miotke

Barducci²,

Astakhova³

2015

Chemosensors

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Detection of low-abundance nucleic acids is a challenging task, which over the last two decades has been solved using enzymatic target amplification. Enzymatic synthesis enhances the signal so that diverse, scientifically and clinically relevant molecules can be identified and studied, including cancer DNA, viral nucleic acids, and regulatory RNAs. However, using enzymes increases the detection time and cost, not to mention the high risk of mistakes with amplification and data alignment. These limitations have stimulated a growing interest in enzyme-free methods within researchers and industry. In this review we discuss recent advances in signal-enhancing approaches aimed at nucleic acid diagnostics that do not require target amplification. Regardless of enzyme usage, signal enhancement is crucial for the reliable detection of nucleic acids at low concentrations. We pay special attention to novel nanomaterials, fluorescence microscopy, and technical advances in detectors for optical assessment. We summarize sensitivity parameters of the currently available assays and devices which makes this review relevant to the broad spectrum of researchers working in fields from biophysics, to engineering, to synthetic biology and bioorganic chemistry.

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Perylene Attached to 2′-Amino-LNA: Synthesis, Incorporation into Oligonucleotides, and Remarkable Fluorescence Properties in Vitro and in Cell Culture

Cited by 52 publications

References 101 publications

Fluorescent oligonucleotides containing a novel perylene 2′-amino-α-L-LNA monomer: Synthesis and analytical potential

Fluorescent oligonucleotides containing a novel perylene 2′-amino-α-L-LNA monomer: Synthesis and analytical potential

Rapid genotyping using pyrene−perylene locked nucleic acid complexes

Novel Signal-Enhancing Approaches for Optical Detection of Nucleic Acids—Going beyond Target Amplification

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