Sulfurizing Reagent Ii and Its Use in Synthesizing Oligonucleotide Phosphorothioates

Preparation of oligoribonucleotides with a phosphorothioate backbone requires a sulfurizing reagent that is efficient and stable in solution. One of the most efficient sulfurizing reagents, 3H1,2-benzodithiol-3-one-1,1-dioxide (Beaucage Reagent), is stable in solution but lacks longterm stability on the DNA synthesizer. This report evaluates Beaucage Reagent and 3((N,N-dimethyl-aminomethylidene)amino)-3H1,2,4-dithiazole-5-thione (DDTT) (Sulfurizing Reagent II) as sulfurizing reagents when using TOM-protected and TBDMS-protected RNA phosphoramidites, and standard DNA phosphoramidites for oligonucleotide synthesis. The product oligonucleotides were evaluated by ion exchange HPLC, ESI-Mass, and 31P NMR. It was confirmed that Ion Exchange HPLC can resolve oligonucleotides containing a single phosphodiester linkage from those containing all phosphorothioate linkages.