- Liquid-phase RNA synthesis by using alkyl-chain-soluble support
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Recent progress in the RNA therapeutics has increased demand for the synthesis of large quantities of oligoribonucleotides. The assembly of RNA oligomers relies mainly on solid-phase approaches. These allow rapid product purification and the ability to drive a target reaction to completion through the use of excess reagents. Despite the known advantages of solid-phase synthesis, some issues in the process remain to be addressed, such as low and limited scale, reagent accessibility, and the use of a very large excess of reagents. Herein, we report a highly efficient and practical method of liquid-phase synthesis of RNA oligomers by using alkyl-chain-soluble support. We demonstrate the utility of the liquid-phase method through 21-mer RNA synthesis on a gram scale. The assembly of RNA oligomers relies principally on solid-phase approaches, although some alternative methods have been developed to date. A highly efficient and practical method of liquid-phase synthesis for RNA oligomers by using an alkyl-chain-type soluble support is reported. The utility of the liquid-phase method through 21-mer RNA synthesis on a gram scale is described (see scheme). Copyright
- Kim, Shokaku,Matsumoto, Masanori,Chiba, Kazuhiro
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supporting information
p. 8615 - 8620
(2013/07/26)
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- Solid-phase synthesis of (poly)phosphorylated nucleosides and conjugates
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Succinyl-cycloSal-phosphate triesters of ribo-and 2′- deoxyribonucleosides were attached to aminomethyl polystyrene as an insoluble solid support and reacted with phosphate-containing nucleophiles yielding nucleoside di-and triphosphates, nucleoside diphosphate sugars, and dinucleoside polyphosphates in high purity after cleavage from the solid support. Here, reactive cycloSal-phosphate triesters were used as immobilized reagents that led to a generally applicable method for the efficient synthesis of phosphorylated biomolecules and phosphate-bridged bioconjugates.
- Tonn, Viktoria Caroline,Meier, Chris
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experimental part
p. 9832 - 9842
(2011/10/09)
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- High yield detritylation of surface-attached nucleosides with photoacid generated in an overlying solid film: Roles of translational diffusion and scavenging
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Conventional solid-phase oligonucleotide synthesis overcomes the reversibility of acid-dependent detritylation by washing away the released dimethoxytrityl cations (DMT+) with acid. This option is unavailable if the acid is photogenerated in an overlying solid film, as in the photolithographic fabrication of oligonucleotide arrays on planar surfaces. To overcome the resulting reversibility problem we developed methods of achieving ≥98% detritylation of glass-attached 5′-O-DMT-thymidine, a model for 5′-O-DMT-protected oligonucleotides, by the photogeneration of trichloroacetic acid in a solid film. Enhanced intrafilm diffusion, insufficient to degrade the photolithographic resolution but enabling DMT+ to move from its plane of release into the overlying photoacid-generating film, increased detritylation from ≤30% to ≥98%. Inclusion of an intrafilm carbocation scavenger such as a triarylsilane hydride converted the detritylation into a time-dependent irreversible process proceeding to ≥99% detritylation within 60 s following brief photoacid generation. Light sensitivity is high, exceeding direct photodeprotection methods by 15-100 fold. The Royal Society of Chemistry 2009.
- Garland, Peter B.,Serafinowski, Pawel J.
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experimental part
p. 451 - 459
(2009/07/18)
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- Solid-phase synthesis of oligonucleotide glycoconjugates bearing three different glycosyl groups: Orthogonally protected bis(hydroxymethyl)-N,N′- bis(3-hydroxypropyl)malondiamide phosphoramidite as key building block
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Diethyl O,O′-(methoxymethylene)bis(hydroxymethyl)malonate (3) was observed to undergo a stepwise aminolysis when treated with 3-aminopropanol. This allowed convenient preparation of bis(hydroxymethyl)-N,N′-bis(3- hydroxypropyl)malondiamide bearing orthogonal levulinyl (Lev) and tert-butyldiphenylsilyl (TBDPS) protections at the two N-hydroxypropyl groups (8). One of the hydroxylmethyl functions was then protected with a 4,4′-dimethoxytrityl (DMTr) group, and the other one was phosphitylated to obtain a methyl N,N-diisopropylphosphoramidite (1). This building block was used for the synthesis of oligonucleotide glycoconjugates (25 and 26) carrying three different sugar units. After conventional phosphoramidite chain assembly of the sequence containing 1, the 5′-terminal DMTr group was removed and an appropriate glycosyl 6-O-phosphoramidite was coupled. The remaining protections of the branching unit were removed in the order of Lev and TBDPS, and the exposed hydroxyl functions were reacted one after another with the desired glycosyl 6-O-phosphoramidites. Global deprotection and cleavage of the conjugate from the support were achieved by conventional ammonolysis.
- Katajisto, Johanna,Heinonen, Petri,Loennberg, Harri
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p. 7609 - 7615
(2007/10/03)
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- Solid Phase Synthesis of Oligodeoxyribonucleotides Utilizing the Phenylthio Group as a Phosphate Protecting Group
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Oligodeoxyribonucleotide synthesis utilizing the phenylthio group as a phosphate protecting group was applied to the solid phase method.The base residues of deoxyguanosine and deoxyadenosine were protected with bis(isobutyryloxy)ethylene (Bibe) and phthaloyl groups to avoid the base modfication and depurination, respectively.A key synthetic intermediate of N2-isobutyryl-N1,N2-bis(isobutyryloxy)ethylenedeoxyguanosine was prepared in high yield by four-step reaction from deoxyguanosine and used for preparation of the building blocks of deoxyguanosine required for the polymer support synthesis.Two kinds of polymer supports, i.e., 1 percent cross-linked polystyrene and controlled pore glass were chosen.The latter was employed for the synthesis of dodecadeoxyribonucleotides by using an automated DNA synthesizer.
- Matsuzaki, Jun-ichi,Kohno, Kyoko,Tahara, Shin-ichiro,Sekine, Mitsuo,Hata, Tsujiaki
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p. 1407 - 1414
(2007/10/02)
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- A comparison of β-functionalized ethyl groups for the protection of the phospho function in decathymidylate synthesis using a phosphite triester approach
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A number of phosphorodichloridites (1) carrying various β-functionalized ethyl protective groups were converted into their corresponding phosphoromorpholido chloridites (2).These were then reacted with 5'-O-(dimethoxytrityl)thymidine to give the pertinent phosphoromorpholidites (3).The applicability of these compounds in the synthesis of oligodeoxynucleotides on a solid support has been evaluated using the synthesis of the thymidine decamer.
- Claesen, C.A.A.,Segers, R.P.A.M.,Tesser, G.I.
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p. 209 - 214
(2007/10/02)
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