Fast Deprotection of Synthetic Oligodeoxyribonucleotides Using Standard Reagents under Microwave Irradiation (original) (raw)
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Microwave-assisted rapid deprotection of oligodeoxyribonucleotides
Nucleic Acids Research, 1997
A novel method for the deprotection of oligodeoxyribonucleotides under microwave irradiation has been developed. The oligodeoxynucleotides having base labile, phenoxyacetyl (pac), protection for exocyclic amino functions were fully deprotected in 0.2 M sodium hydroxide (methanol:water : : 1:1, v/v) = A and 1 M sodium hydroxide (methanol:water : : 1:1, v/v) = B using microwaves in 4 and 2 min, respectively. The deprotection of oligodeoxyribonucleotides carrying conventional protecting groups, dA bz , dC bz and dG pac , for exocyclic amino functions was achieved in 4 min in B without any side product formation. The deprotected oligonucleotides were compared with the oligomers deprotected using standard deprotection conditions (29% aq. ammonia, 16 h, 55_C) with respect to their retention time on HPLC and biological activity.
Advanced method for oligonucleotide deprotection
Nucleic Acids Research, 2000
A new procedure for rapid deprotection of synthetic oligodeoxynucleotides has been developed. While all known deprotection methods require purification to remove the residual protective groups (e.g. benzamide) and insoluble silicates, the new procedure based on the use of an ammonia-free reagent mixture allows one to avoid the additional purification steps. The method can be applied to deprotect the oligodeoxynucleotides synthesized by using the standard protected nucleoside phosphoramidites dG iBu , dC Bz and dA Bz .
Nucleic Acids Research, 1986
3-Methoxy-4-phenoxybenzoyl group has been used for the protection of exocyclic amino group of nucleosides. In case of 2'deoxycytidine it has been found to be highly selective under controlled conditions. The N-protected derivatives of 2'-deoxyadenosine and 2 '-deoxyguanosine have been found to be suf ficiently stable towards acids minimising depurination under conditions of synthesis of oligodeoxyribonucleotide on solid support via phosphotriester approach. The high lipophilicity of the group and milder deprotection conditions are additional advantages.
Deprotection of methylphosphonate oligonucleotides using a novel one-pot procedure
Nucleic Acids Research, 1993
or T were treated with ethylenediamine. A novel method of deprotecting methylphosphonate oligonucleotides was developed from these studies. The method incorporates a brief treatment with dilute ammonia for 30 minutes followed by addition of ethylenediamine for 6 hours at room temperature to complete deprotection in a one-pot format. The solution Is then diluted and neutralized to stop the reaction and prepare the crude product for chromatographlc purification. This method was used to successfully deprotect a series of oligonucleotides at the 1, 100, and 150 /imole scales. These deprotection results were compared to a commonly used two-step method and found to be superior In yield of product by as much as 250%.
Nucleic Acids Research, 1977
Nucleoside 3'-phosphotriesters as key intermediates for the oligoribonucleotide synthesis. IV. New method for removal of 2,2,2-trichloroethyl group and 31P NM R as a new tool for analysis of deblocking of intemucleotide phosphate protecting groups1 ABSTRACT Zinc/acetylacetone/pyridine treatment has been designed as a very efficient method for removal of 2,2,2-trichloroethyl group from phosphoesters. Internucleotide and terminal 2,2,2-trichloroethylphosphotriesters were transformed to corresponding diesters quantitatively. Much less reactive 2,2,2-trichloroethylphosphodiesters produced monoesters with ca. 909 yield. 31 NMR spectroscopy has been proposed as a new tool for analysis of removal of internucleotide phosphate protecting groupsa crucial step in oligonucleotides synthesis via phosphotriester approach.
Journal of Organic Chemistry, 2008
Emerging RNA-based technologies for controlling gene expression have triggered a high demand for synthetic oligoribonucleotides and have motivated the development of ribonucleoside phosphoramidites that would exhibit coupling kinetics and coupling efficiencies comparable to those of deoxyribonucleoside phosphoramidites. To fulfill these needs, the novel 4-(N-dichloroacetyl-N-methylamino)benzyloxymethyl group for 2′-hydroxyl protection of ribonucleoside phosphoramidites 9a-d has been implemented (Schemes 1 and 2). The solid-phase synthesis of AUCCGUAGCUAACGUCAUGG was then carried out employing 9a-d as 0.2 M solutions in dry MeCN and 5-benzylthio-1H-tetrazole as an activator. The coupling efficiency of 9a-d averaged 99% within a coupling time of 180 s. Following removal of all basesensitive protecting groups, cleavage of the remaining 2′-[4-(N-methylamino)benzyl] acetals from the RNA oligonucleotide was effected in buffered 0.1 M AcOH (pH 3.8) within 30 min at 90°C. RP-HPLC and PAGE analyses of the fully deprotected AUCCGUAGCUAACGUCAUGG were comparable to those of a commercial RNA oligonucleotide sharing an identical sequence. Enzymatic digestion of the RNA oligomer catalyzed by bovine spleen phosphodiesterase and bacterial alkaline phosphatase revealed no significant amounts of RNA fragments containing (2′f5′)-internucleotidic phosphodiester linkages or noteworthy nucleobase modifications.
Further Optimization of Detritylation in Solid-Phase Oligodeoxyribonucleotide Synthesis
Nucleosides, Nucleotides and Nucleic Acids, 2011
Various conditions for optimum detritylation (i.e., the removal of 5-O-trityl protecting groups) during solid-phase synthesis of oligodeoxyribonucleotides were investigated. Di-and tri-chloroacetic acids of variable concentrations were used to study the removal of the 4,4-dimethoxytrityl (DMTr) group. It was found that the DMTr group could be completely removed under much milder acidic conditions than what are currently used for automated solid-phase synthesis. The 2,7-dimethylpixyl (DMPx) is proposed as an alternative and more readily removable group for the protection of the 5-OH functions both in solid-and solution-phase synthesis. The improved detritylation conditions are expected to minimize the waste and offer a protocol for incorporation of acid sensitive building-blocks in oligonucleotides.
Nucleic Acids Research, 1999
Two sets of deprotection conditions have been evolved for the deprotection of oligodeoxyribonucleotides and their cleavage from commercially available cis-diol group-bearing universal polymer supports. In the first case, oligodeoxyribonucleotides anchored on the universal support were subjected to one of the standard deprotection conditions followed by treatment with aqueous 0.5 M sodium chloride + 0.2 M sodium hydroxide solution for 30 min at room temperature. In the second case, oligonucleotides bound to the universal support were treated with methanolic sodium hydroxide solution under microwave radiation to obtain fully deprotected oligomers within 4 min. Under both conditions, the cleavage of oligonucleotides from the support and their deprotection occurred quantitatively without any side product formation. The cleaved oligonucleotides were found to be identical in all respects (retention time on HPLC and biological activity in PCR) to the corresponding standard oligonucleotides.