Fast oligonucleotide deprotection phosphoramidite chemistry for DNA synthesis (original) (raw)
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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, 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.
Selective O-phosphitilation with nucleoside phosphoramidite reagents
Nucleic Acids Research, 1992
In contrast to tetrazole, pyridine hydrochloride/ imidazole converts nucleoside phosphoramidltes to intermediates that show a high preference for phosphltllating hydroxyl groups relative to nucleoside amino groups. Use of this activating agent and incorporation of a pyridine hydrochloride/aniline wash step in the synthetic cycles permit synthesis of mixed base twenty-mer oligonucleotides from nucleoside reagents containing unprotected amino groups. This approach should be useful for the synthesis of oligonucleotide analogues containing substltuents sensitive to reagents used in conventional deblocking steps. Pyridine hydrochloride itself is an effective reagent for activating nucleoside methylphosphonoamidites and ribonucleoside phosphoramidltes, as well as deoxyribonucleoside phosphoramldites, when high O/N selectively is not needed.
Nucleic Acids Research, 1976
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.
Synthesis of Protected Amino Hexitol Nucleosides as Building Blocks for Oligonucleotide Synthesis
Journal of Organic Chemistry, 2018
A new synthesis protocol for the preparation of hitherto unknown 1′,5′-anhydro-4′-amino-trityl/MMTr hexitol nucleosides has been developed. Key steps in the synthesis of the pyrimidine analogues (U and C) include the regioselective D-allo-hexitol oxirane and 2′,4′-anhydronucleoside ring opening by uracil and azide, respectively. A different strategy using a regioselective epoxide ring opening of D-gulooxirane, followed by a S N 2 type of azidation reaction, has been adopted for the purine analogues (A and G). These compounds can be easily converted to 6′-phosphoramidites for the solid-phase synthesis of N4′ → P6′ phosphoramidates of amino hexitol nucleic acids (AHNA).
Nucleic Acids Research, 1986
Phosphoramidite reagents can phosphitylate guanine bases at the O-position during solid phase synthesis and serious chain cleavage occurs if the base phosphitylation is not eliminated before the iodine/water oxidation step. This can be accomplished by i) blocking the O^-position with a 2-cyanoethyl protecting group for deoxyribonucleotides or with a p-nitrophenylethyl group for ribonucleotides, ii) regenerating the guanine base with water or acetate ions, or iii) using N-methylanilinium trifluoroacetate (TAMA) as the phosphoramidite activator. The effectiveness of these methods was demonstrated by both 3 1 P NMR studies and by the synthesis of d(Gp) 2 3G, (Gp) 14 G, and d-(Gp) 1 3rG sequences.
Oligonucleotides, 2008
Fast methods for the removal of permanent amide exo-cyclic protective groups widely used in phosphoramidite-method DNA synthesis are desirable for many genomics and proteomics applications. In this communication, we present a method for the deprotection of a range of N-acyl deoxyribonucleosides (T, dA Bz , dC Bz , dC Ac , dG ibu , dG PAC ) and synthetic oligodeoxyribonucleotides, ranging in length from 5-mer to 50-mer. Oligodeoxyribonucleotides were synthesized using standard amide protecting groups (dA Bz , dC Bz , dG ibu ) and phosphoramidite chemistry on cis-diol solid phase support. This deprotection method utilizes 29% aqueous ammonia solution at 170°C for 5 minutes under monomode microwave irradiation at a 20-nmole reaction scale. Reaction products were analyzed by TLC, RP-HPLC, CE, ESI-MS, real-time PCR, agarose gel electrophoresis, and by DNA uracil glycosylase (UDG) and phosphodiesterase I (PDE) enzymatic digestions.
Phosphotriester approach to the synthesis of oligonucleotides: a reappraisal
Journal of the Chemical Society, Perkin Transactions 1, 1993
The phosphotriester approach to the synthesis of oligodeoxyribo-and oligoribo-nucleotides in solution has been reinvestigated. The efficacy of mesitylene-2-sulfonyl chloride (MSCI) 15a, 2,4,6triisopropylbenzenesulfonyl chloride (TrisCI) 1 5b, 4bromobenzenesulfonyl chloride 1 5c. naphthalene-1-sulfonyl chloride 39, and 2-and 4-nitrobenzenesulfonyl chlorides 40a and 40b, respectively, as activating agents has been examined. The latter arenesulfonyl chlorides have been used in conjunction with the following nucleophilic catalysts: 1-methylimidazole, 3-nitro-I H-1,2,4-triazole 19, 5-(3-nitrophenyl)-l H-tetrazole 20a, 5-(3.5-dinitrophenyl)-l H-tetrazole 20b. 5-(1-methylimidazol-2y l)-I H-tetrazole 21, 5-[ (1-methylimidazol-2-yl)methyl]-1 H-tetrazole 22, 4-ethoxypyridine 1-oxide 14a. 4,6-dinitro-l-hydroxybenzotriazole 29a, 1-hydroxy-4-nitro-6-(trifluoromethyl) benzotriazole 29b. 1-hydroxy-5-phenyltetrazole 30a and 1-hydroxy-5-(3-nitrophenyl) tetrazole 30b. The rates of formation and yields of the fully protected dideoxyribonucleoside and diribonucleoside phosphates 37 and 47, respectively, were determined using various combinations of activating agents and nucleophilic catalysts. Although 2-and 4-nitrobenzenesulfonyl chlorides 40a and 40b. respectively, proved to be the most powerful activating agents, their use in the deoxy-series led to the formation of by-products and hence to unsatisfactory isolated yields of the dideoxyribonucleoside phosphate 37. I I +-0 ii. iii /o 3 4 Ar = 2-chlorophenyl; B and B' are protected in substrates 1, 2 and 3, and unprotected base residues in product 4