LARGE-SCALE SYNTHESIS, PURIFICATION, AND ANALYSIS OF OLIGODEOXYNUCLEOTIDE PHOSPHOROTHIOATES

Synthesis of oligonucleotides has been carried out on 1-, 2-, and 5-mm ol scales using an appropriately modified automated, commercially avai lable DNA synthesizer. The reaction cycles were optimized to obtain ef ficient coupling (greater than or equal to 97%). The synthesized oligo nucleotide was purified by preparative reversed-phase liquid chromatog raphy, followed by detritylation and desalting to obtain the oligonucl eotides in the Na+ form. The purified oligonucleotides were characteri zed by P-31 NMR, mass spectrometry, capillary gel electrophoresis, and ion-exchange high-performance liquid chromatography. By using these p rotocols, a 25-mer oligodeoxyribonucleotide can be synthesized on a 1- , 2-, or 5-mmole scale to obtain approximately 2.4, 4.8, or 12 g of pu rified product.