RESUMEN
31P cross polarization (CP) magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra were acquired for various linear and branched di- and tri-nucleotides attached to a controlled pore glass (CPG) solid support. The technique readily distinguishes the oxidation state of the phosphorus atom (phosphate versus phosphate), the presence or absence of a protecting group attached directly to phosphorus (cyanoethyl), and other large changes in the phosphorus chemistry (phosphate versus phosphorothioate). However, differences in configurational details remote from the phosphorus atom, such as the attachment position of the ribose sugar (2'5' versus 3'5'), or the particulars of the nucleotide bases (adenine versus uridine versus thymine), could not be resolved. When different stages of the oligonucleotide synthetic cycle were examined, 31P CPMAS NMR revealed that the cyanoethyl protecting group is removed during the course of chain assembly.
Asunto(s)
Espectroscopía de Resonancia Magnética/métodos , Oligonucleótidos/química , Fósforo/química , Química Orgánica/métodos , Vidrio/química , Isótopos de FósforoRESUMEN
The chemical synthesis of oligoribonucleotides containing vicinal (2'-5')- and (3'-5')-phosphodiester linkages is described. The solid-phase method, based on silyl-phosphoramidite chemistry, was applied to the synthesis of a series of branched RNA [(Xp)nA2' (pN)n3'(pN)n] related to the splicing intermediates derived from Saccharomyces cerevisiae rp51a pre-messenger RNA. The branched oligonucleotides have been thoroughly characterized by nucleoside and branched nucleotide composition analysis. Branched oligoribonucleotides will be useful in the study of messenger RNA splicing and in determining the biological role of RNA 'lariats' and 'forks' in vivo.
Asunto(s)
Oligorribonucleótidos/síntesis química , Empalme del ARN , ARN Mensajero/síntesis química , Secuencia de Bases , Cromatografía en Gel , Cromatografía Líquida de Alta Presión , Electroforesis en Gel de Poliacrilamida , Indicadores y Reactivos , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Oligorribonucleótidos/química , Oligorribonucleótidos/aislamiento & purificación , Precursores del ARN/síntesis química , Precursores del ARN/químicaAsunto(s)
Oligonucleótidos/química , Secuencia de Bases , Datos de Secuencia Molecular , Conformación de Ácido Nucleico , Oligodesoxirribonucleótidos/síntesis química , Oligodesoxirribonucleótidos/química , Oligonucleótidos/síntesis química , Oligorribonucleótidos/síntesis química , Oligorribonucleótidos/química , Precursores del ARN/genética , Precursores del ARN/metabolismo , Empalme del ARN , Saccharomyces cerevisiae/genéticaRESUMEN
A simplified and economical method for the attachment of 2'-deoxyribo, ribo and arabinonucleosides onto long-chain alkylamidopropanoic acid controlled-pore glass (LCAAP-CPG, P-3) is described. In this procedure, 5'-O-tritylated nucleosides are coupled directly to LCAAP-CPG in excellent yields using 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (DEC) as coupling reagent. The conventional and time-consuming preparation of nucleoside-3'-O-succinates is no longer required.