SpringerProtocols: Abstract: RNA Synthesis Using 2′-O-(Tert-Butyldimethylsilyl) Protection

RNA Synthesis Using 2′-O-(Tert-Butyldimethylsilyl) Protection

By: Brian S. Sproat²

Abstract

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This chapter enables the reader to carry out the solid-phase synthesis of ribonucleic acid (RNA) using β-cyanoethyl phosphoramidite chemistry combined with tert-butyldimethylsilyl protection of the ribose 2′-hydroxyl group. Phosphoramidite monomers are activated with 5-benzylmercapto-1H-tetrazole enabling fast and highly efficient coupling to the 5′-hydroxyl group of the support-bound oligonucleotide. On completion of the synthesis, the stepwise deprotection of the nucleobase, phosphate, and ribose protecting groups is carried out using optimized protocols. Subsequently the various high-pressure (performance) liquid chromatography (HPLC) procedures are described enabling the purification and analysis of the RNA. For this purpose anion-exchange and reversed-phase HPLC are used singly or in combination according to the final purity requirement of the RNA.

Affiliation(s): (2) RNA-TEC, Leuven, Belgium

Book Title: Oligonucleotide Synthesis

Series: Methods in Molecular Biology | Volume: 288 | Pub. Date: Sep-01-2004 | Page Range: 17-31 | DOI: 10.1385/1-59259-823-4:017

Subject: Biochemistry

Key Words: Oligoribonucleotide synthesis - TBDMS - 5-benzylmercapto-1H-tetrazole - solid phase - triethylamine tris(hydrofluoride) - anion exchange - HPLC - β-cyanoethyl phosphoramidite - chaotropes

References

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