RNA synthesis using 2'-O-(tert-butyldimethylsilyl) protection.

This chapter enables the reader to carry out the solid-phase synthesis of ribonucleic acid (RNA) using beta-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.