Oligonucleotides Containing N7-Cyanoborane-2′-deoxyguanosine

Abstract

For synthesis of N⁷-cyanoborane-containing oligonucleotides, the 5′-DMT protecting group is not a suitable precursor because the boronated nucleoside is incompatible with DMT cations released during deprotection of the oligonucleotide. As an alternative to DMT, we have investigated use of the 5′-Fmoc protecting group. We found that the cyanoborane group is stable during synthesis and deprotection conditions used with Fmoc derivatives.     

Rapid Purification of Chemically Synthesized Oligodeoxy-Nucleotides

Abstract

Advances in solid phase oligonucleotide synthesis have increased th number of sequences that can be synthesized within a short time. However, various deprotection steps are required for the isolation of the final products in a pure form. These work-up procedures are more time consuming than the synthesis itself.     

Reusable Solid-Phase Supports for Oligonucleotide Synthesis Using Hydroquinone-O,O'-diacetic Acid (“Q-Linker”)

Abstract

Reusable solid-phase supports for large scale oligonucleotide synthesis have been prepared by converting amino derivatized supports into hydroxyl supports. Rapid nucleo side attachment, via a Q-linker arm, was automatically performed on the DNA synthesizer using HBTU and DMAP as the coupling reagents. All steps were suitable for automation and ～ 1.5 h was required to prepare the supports for reuse. Up to twelve consecutive syntheses of a 20-mer phosphorothioate were performed on a synthesis column.     

Structural Determination of Silicon-Containing Oligonucleotides by 1H−29Si Long-Range Heteronuclear Multiple Quantum Correlation NMR Spectroscopy

Abstract

The technique of ¹H^?29Si Long-Range Heteronuclear Multiple Quantum Correlation NMR Spectroscopy was used to determine the structure of silicon-containing oligonucleotides. Trimers which contained silicon instead of phosphorus as part of the oligonucleotide link were synthesized through a synthetic route that required minimal hydroxyl protection. The resulting trimer could have one of two possible structures. Through the use of ¹H^?29Si HMQC NMR spectroscopy, it was possible to link the 3′-hydroxymethine proton of one sugar to the 5′-hydroxymethylene proton of an adjacent sugar by correlation to the same silicon atom, thus elucidating the final structure.     

6-AZAPYRIMIDINE AND 7-DEAZAPURINE 2′-DEOXY-2′-FLUOROARABINONUCLEOSIDES: SYNTHESIS,CONFORMATION AND PROPERTIES OF OLIGONUCLEOTIDES

The synthesis of 2′-deoxy-2′-fluoro-β-d-arabinofuranosyl nucleosides (1b, 2b, and 3b) were described and their conformation in solution as well as in the solid state was determined. In addition to this, building blocks 10a,b and 13a,b were prepared and employed in solid-phase oligonucleotide synthesis. For compounds 1a and 1b the lactime proton is protected to avoid unresolved degradation of its phosphoramidites 10a,b. UV-melting studies have been carried out to assess the thermal stability of oligonucleotides containing compounds 1a,b, and 3a,b.     

Oligonucleotides Containing Pyrazolo[3,4-d]Pyrimidines: 8-Aza-7-deazaadenines With Bulky Substituents in the 2- or 7-Position

The synthesis of the 2′-deoxyadenosine analogues 1b, 2b, and 3c modified at the 7- and/or 2-position is described. The effect of 7-chloro and 2-methylthio groups on the duplex stability is evaluated. For that, the nucleosides 1b, 2b, and 3c were converted to the corresponding phosphoramidites 15, 19, and 22, which were employed in the solid-phase oligonucleotide synthesis. In oligonucleotide duplexes, compound 1b forms stable base pairs with dT, of which the separated 1b- dT base pairs contribute stronger than that of the consecutive base pairs. Compound 2b shows universal base pairing properties while its N8 isomer 3c forms duplexes with lower stability.     

Etude des Parametres Experimentax Influencant la Formation de la Liaison Phosphotriester Activee par le BOP. Application en Synthese Oligodesoxyribonucleotidique

Abstract

The optimum conditions for the use of BOP as the condensing reagent in oligonucleotide synthesis have been determinated. They were applied to a rapid preparation of an undecanucleoside decaphosphate.     

NMR Studies of the Interaction of Bleomycin with (dC-dG)3

Abstract

The interaction of bleomycin A₂ and Zn(II)-bleomycin A₂ with the oligonucleotide (dC-dG)₃ has been monitored by nuclear magnetic resonance spectroscopy. Binding of the drug to the oligonucleotide is indicated by an upfield shift of the bithiazole proton resonances consistent with partial intercalation of this group between base pairs. The effect of temperature and ionic strength on the binding of both free bleomycin and the Zn(II) complex has been studied. Consistent with earlier studies on polynucleotides, the rate of exchange between the free drug and the drug-oligonucleotide complex is rapid on the ¹H NMR chemical shift time scale. Binding of the oligonucleotide induced changes in resonances assigned to protons in the metal-binding region of Zn(II)-bleomycin. Intermolecular nuclear Overhauser effect enhancements between bleomycin and the oligonucleotide have not been detected.     

Chemical and Photochemical Ligation of Oligonucleotide Blocks

Abstract

Several efficient means for joining oligonucleotides in dilute solution by non-natural internucleotide bridges are discussed. It is also shown that an oligonucleotide containing a -OP(O)(O^?)S- link can hnction as an effective template in PCR amplification and that oligonucleotide probes containing stilbenedicarboxamide groups can serve in monitoring the presence of mismatched bases in an oligonucleotide target.     

Covalent Attachment of Methylene Blue to Oligonucleotides

Abstract

The synthesis and application of oligonucleotides derivatized by methylene blue are described. For that, a carboxylated methylene blue derivative was synthesized and transformed into an activated N-hydroxysuccinimidoester. The activated ester was reacted with 5′-aminoalkylated oligonucleotides. The labelled oligonucleotides were isolated and characterized both by reversed phase HPLC and MALDI mass spectrometry. Initial studies on analytical application of these oligonucleotide conjugates are discussed.     

Evaluation of Sulfur-Transfer Reagents for Large Scale Synthesis of Oligonucleotide Phosphorothioate Analogues

Abstract

Several sulfur-transfer reagents have been evaluated for large scale synthesis of oligonucleotide phosphorothioate analogues, in which 3-ethoxy-1, 2-dithiazoline-5-one (EDITH, 5) shows potential as an alternative to Beaucage reagent.     

Selective Attachment of Oligonucleotides to Interleukin-1β and Targeted Delivery to Cells

Abstract

Conjugates between oligodeoxyribonucleotides and an interleukin-1β mutant protein have been constructed using a heterobifunctional cross-linker. These protein-DNA conjugates had conserved binding activity to the interleukin-1 receptor. The oligonucleotide hybridization properties were unchanged.     

Oligonucleotide Labeling Methods 4. Direct Labeling Reagents with a Novel,Non-Nucleosidic,Chirally Defined 2-Deoxy-β-D-Ribosyl Backbone.

Abstract

Novel solid supports and CE-phosphoramidite reagents have been prepared featuring a unique 2′-deoxyribosyl backbone. These chirally pure reagents form the basis of an oligonucleotide labeling system which provides diastereomerically pure oligonucleotides.     

OLigonucleotide (Cyanomethyl) Phosphonates

Abstract

A dinucleoside (cyanomethyl) phosphonate has been prepared, and its properties have been studied. This compound was converted into an oligonucleotide possessing alternating (cyanomethyl) phosphonate and phosphodiester backbone groups and its hybridization to complementary DNA and RNA sequences was studied versus methylphosphonate and phosphodiester controls.     

Triple Helix Structures: Sequence Dependence,Flexibility and Mismatch Effects

Abstract

By means of molecular modelling, electrostatic interactions are shown to play an important role in the sequence-dependent structure of triple helices formed by a homopyrimidine oligonucleotide bound to a homopurine, homopyrimidine sequence on DNA. This is caused by the presence of positive charges due to the protonation of cytosines in the Hoogsteen-bonded strand, required in order to form C.GxC+ triplets. Energetic and conformational characteristics of triple helices with different sequences are analyzed and discussed. The effects of duplex mismatches on the triple helix stability are investigated via thermal dissociation using UV absorption.     

Triplex Formation Involving 2′-O,4′-C-Methylene Bridged Nucleic Acid (2′,4′-BNA) with 2-Pyridone Base Analogue: Efficient and Selective Recognition of C:G Interruption

Abstract

For the effective recognition of C:G interruption in homopurine-homopyrimidine duplex DNA, we examined triplex-forming ability and sequence-selectivity of a triplex-forming oligonucleotide (TFO) involving of 2′-O,4′-C-methylene bridged nucleic acid with 2-pyridone base analogue. We found that the modified TFO formed stable triplex with high binding affinity and sequence-selectivity.     

Synthesis and Binding Properties of Oligonucleotides Containing an Azobenzene Linker

Abstract

Incorporation of an azobenzene-4,4′-diamide group via a linker arm into the 3′-hydroxyl function of one oligonucleotide segment and the 5′-OH of other oligonucleotide has been described. The binding of the oligonucleotides containing the azobenzene linker was investigated by UV melting behaviors. The azobenzene linker has been shown to be useful as an effective bridge for stabilizing hairpin duplex and triplex.     

Triplex Formation Involving 2′-O,4′-C-Methylene Bridged Nucleic Acid (2′,4′-BNA) with 1-Is oquinolone Base Analogue: Efficient and Selective Recognition of C:G Interruption

Abstract

For the effective recognition of C ? G interruption in homopurine-homopyrimidine duplex DNA, we examined triplex-forming ability and sequence-selectivity of a triplex-forming oligonucleotide (TFO) involving of 2′-O, 4′-C-methylene bridged nucleic acid with 1-isoquinolone base analogue. We found that the modified TFO formed stable triplex with high binding affinity and sequence-selectivity.     

Recognition of Point Mutations in Dna By Means of Time Resolved Fluorescence Methods

Abstract

A rapid recognition of aberrations in the base sequence of nucleic acids is an important step toward the diagnosis of genetic diseases. We have developed a hybridization method in which a fluorescently labeled oligonucleotide is used to detect point mutations in a target by a simple fluorescence lifetime analysis of the emission of the fluorescent label.     

Rational Methods of DNA Synthesis,Exemplified for the Preparation of Parts of the Human Proopiomelanocortin Gene

Abstract

Complete genes can now be prepared in single-batch procedures a) as one long oligonucleotide strand, b) as a number of simultaneously synthesized fragments. For further increase of overall efficiency DNA fragments terminated by a 3′-ribonucleoside can be joined by solid-phase single strand ligation using RNA ligase. This paper describes the application of these techniques to the preparation of parts of the human proopiomelanocortin gene.