Développement de nouvelles sondes oligonucléotidiques fluorescentes pour l'analyse des acides nucléiques

Thèse financée par la Région CentreNew fluorescent oligonucleotide probes for the specific recognition of double-stranded nucleic acid structures were obtained. These triple helix forming oligonucleotides were linked through their 5'-or/and 3'-ends to a series of cyanine monomethine labels, with intercalating properties, via polymethylene linkers. In all cases, the hybridization of the labelled oligonucleotides with their double-stranded targets induced a strong increase of the fluorescence emission. The fluorescence increase was dependent on both the cyanine and the parameters of its linkage to the oligonucleotides (linker size and positions of its attachment to both the label and the oligonucleotide). In the presence of the single-stranded target, the fluorescence increase was weak. The triplex structures were stabilized by the presence of the labels. The probes involving thiazole orange attached by the benzothiazole ring and an octamethylene link provided the best balanced properties in terms of triplex stabilization, fluorescence intensity and fluorescence enhancement upon hybridization with the double-stranded target.With the aim to obtain new probes that can be used in cellular and in vivo experiments, squaraine dyes were synthesized and used, for the first time to our knowledge, for the labelling of oligonucleotides. These squaraine-oligonucleotide conjugates are detectable beyond 650 nm and possess good quantum yields. The photophysical properties of the conjugates at 37 °C and in moderate acidic conditions can be strongly improved by the incorporation of fluorine atoms on the squaraine dye.De nouvelles sondes oligonucléotidiques fluorescentes spécifiques des structures d'acides nucléiques en double brin ont été obtenues. Ces oligonucléotides formant des triples hélices, sont liés par leurs extrémités 5'-ou/et 3' à une série de marqueurs fluorescents de la famille des cyanines monométhines possédant des propriétés intercalantes, par des bras polyméthylène de longueur variable. Dans tous les cas, l'hybridation des sondes avec les cibles en double brin s'accompagne d'une forte augmentation de l'intensité du signal fluorescent émis qui dépend de la structure de la cyanine et des paramètres de sa liaison aux oligonucléotides (longueur du bras de liaison et sa position de fixation sur les oligonucléotides et sur les marqueurs). En présence d'une cible simple brin, les modifications du signal ne sont pas significatives. La présence du marqueur stabilise les triples hélices. Les sondes comportant le thiazole orange, attaché par le noyau benzothiazole via un bras de liaison à huit méthylènes, présente le meilleur compromis en termes de stabilisation, d'intensité de fluorescence et d'augmentation d'intensité de fluorescence lors de l'hybridation.Dans le but d'obtenir de nouvelles sondes utilisables en milieu cellulaire et in vivo, de nouveaux marqueurs fluorescents de type squaraine ont été synthétisés et couplés pour la première fois, à notre connaissance, à des oligonucléotides. Les conjugués correspondant émettent un signal fluorescent détectable au-delà de 650 nm et possèdent des rendements quantiques de fluorescence élevés. La photostabilité des marqueurs et des conjugués, en milieu acide faible et à 37 °C a été fortement augmentée après incorporation d'atomes de fluor sur le squaraine

A simple synthesis of 2-keto-3-deoxy-D-erythro-hexonic acid isopropyl ester, a key sugar for the bacterial population living under metallic stress

International audience2-Keto-3-deoxy-D-erythro-hexonic acid (KDG) is the key intermediate metabolite of the Entner Doudoroff (ED) pathway. A simple, efficient and stereoselective synthesis of KDG isopropyl ester is described in five steps from 2,3-O-isopropylidene-D-threitol with an overall yield of 47%. KDG isopropyl ester is studied as an attractive marker of a functional Entner Doudoroff pathway. KDG isopropyl ester is used to promote growth of ammonium producing bacterial strains, showing interesting features in the remediation of heavy-metal polluted soils

Lewis acid catalysis and Green oxidations: sequential tandem oxidation processes induced by Mn-hyperaccumulating plants

International audienceAmong the phytotechnologies used for the reclamation of degraded mining sites, phytoextraction aims to diminish the concentration of polluting elements in contaminated soils. However, the biomass resulting from the phytoextraction processes (highly enriched in polluting elements) is too often considered as a problematic waste. The manganese-enriched biomass derived from native Mn-hyperaccumulating plants of New Caledonia was presented here as a valuable source of metallic elements of high interest in chemical catalysis. The preparation of the catalyst Eco-Mn1 and reagent Eco-Mn2 derived from Grevillea exul exul and Grevillea exul rubiginosa was investigated. Their unusual polymetallic compositions allowed to explore new reactivity of low oxidative state of manganese-Mn(II) for Eco-Mn1 and Mn(IV) for Eco-Mn2. Eco-Mn1 was used as a Lewis acid to catalyze the acetalization/elimination of aldehydes into enol ethers with high yields; a new green and stereoselective synthesis of (-)-isopulegol via the carbonyl-ene cyclization of (+)-citronellal was also performed with Eco-Mn1. Eco-Mn2 was used as a mild oxidative reagent and controlled the oxidation of aliphatic alcohols into aldehydes with quantitative yields. Oxidative cleavage was interestingly noticed when Eco-Mn2 was used in the presence of a polyol. Eco-Mn2 allowed direct oxidative iodination of ketones without using iodine, which is strongly discouraged by new environmental legislations. Finally, the combination of the properties in the Eco-Mn catalysts and reagents gave them an unprecedented potential to perform sequential tandem oxidation processes through new green syntheses of p-cymene from (-)-isopulegol and (+)-citronellal; and a new green synthesis of functionalized pyridines by in situ oxidation of 1,4-dihydropyridines

Fluorinated squaraine as near-IR label with improved properties for the labeling of oligonucleotides

International audienceA new squaraine dye with fluorinated benzothiazole rings was synthesized. This new label possesses improved photophysical properties and chemical stability as compared to the corresponding non-fluorinated and the dicyanosquaraines. These squaraines were used for the labeling of a series of oligonucleotides with various sequences, lengths, and chemistries. The conjugates involving the fluorinated squaraine possess the best properties: emission wavelength >670 nm, high quantum yields (0.27-0.39)

Targeting DNA with "light-up" pyrimidine triple-helical forming oligonucleotides conjugated to stabilizing fluorophores

International audienceThe synthesis of triple-helix-forming oligonucleotides (TFOs) linked to a series of cyanine monomethines has been performed. Eight cyanines including one thiocyanine, four thiazole orange analogues, and three quinocyanines were attached to the 5'-end of 10-mer pyrimidine TFOs. The binding properties of these modified TFOs with their double-stranded DNA target were studied by absorption and steady-state fluorescence spectroscopy. The stability of the triplex structures depended on the cyanine structure and the linker size used to connect both entities. The most efficient cyanines able to stabilize the triplex structures, when attached at the 5'-end of the TFO, have been incorporated at both ends and provided triplex structures with increased stability. Fluorescence studies have shown that for the TFOs involving one cyanine, an important intensity increase (up to 37-fold) in the fluorescent signal was observed upon their hybridization with the double-stranded target, proving hybridization. The conjugates involving thiazole orange attached by the benzothiazole ring provided the most balanced properties in terms of triplex stabilization, fluorescence intensity and fluorescence enhancement upon hybridization with the double-stranded target. In order to test the influence of different parameters such as the TFO sequence and length, thiazole orange was used to label 17-mer TFOs. Hybridizations of these TFOs with different duplexes, designed to study the influence of mismatches at both internal and terminal positions on the triplex structures, confirmed the possibility of triplex formation without loss of specificity together with a strong fluorescence enhancement (up to 13-fold)

USES OF CERTAIN PLATINOID ACCUMULATING PLANTS FOR USE IN ORGANIC CHEMICAL REACTIONS

The invention relates to the use of products derived from ash from plants that accumulate metals from the platinum group (platinoids) for use in chemical reactions

USES OF CERTAIN PLATINOID ACCUMULATING PLANTS FOR USE IN ORGANIC CHEMICAL REACTIONS

The invention relates to the use of products derived from ash from plants that accumulate metals from the platinum group (platinoids) for use in chemical reactions

Stabilization of hairpins and bulged secondary structures of nucleic acids by single incorporation of α,β-D-CNA featuring a gauche(+) alpha torsional angle

A constrained dinucleotide unit featuring a gauche(+) alpha torsional angle configuration was used to stabilize DNA hairpin or bulged structures. Large five nucleotides having looped hairpin structures can be stabilized up to +5 degrees C. Depending on the nature of the closing base pair, the increased hairpin stability can be reached through loop moiety preorganization or stem rearrangement induced on the first two base pairs. With an alpha gauche(+) constraint either within or opposite to the bulge, the larger bulged structures had better stabilization

C-C bond formation strategy through palladium Ecocatalysis: from palladium recycling to innovative catalysts

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C-C bond formation strategy through ecocatalysis: Insights from structural studies and synthetic potential

International audienceWe report the elaboration of novel bio-sourced ecocatalysts for Heck and Suzuki cross-coupling reactions.Ecocatalysis is based on the recycling of metals issued from phytoremediation or rehabilitation, and aninnovative chemical valorization of the subsequent biomass in the field of catalysis. Here, we describe theefficient palladium accumulation by plants (Brassica juncea, Lolium multiflorum) via rhizofiltration. Takingadvantage of the remarkable ability of the selected plants to accumulate Pd(II) species into their roots,these latter can be directly used for the preparation of ecocatalysts, called Eco-Pd®. The formed Eco-Pd®catalysts are thoroughly characterized via ICP-MS, XRD, XPS, TEM, SEM in order to elucidate the chemicalcomposition and morphology of the formed materials. Significant differences to conventional Pd-basedcatalysts such as palladium(II) chloride can principally be related to a particular Lewis acid behavior ofthe Eco-Pd®catalysts. Finally, the obtained Eco-Pd®appear as highly active catalysts in Heck and Suzukicross-coupling reactions necessitating considerably lower Pd quantity compared to precedently reportedbio-sourced palladium containing catalysts