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Updated: Sep 30, 2026

Chemical Triphosphorylation of Oligonucleotides
Published on: June 2, 2022
Separation of synthetic oligonucleotide dithioates from monothiophosphate impurities by anion-exchange chromatography
Xianbin Yang1, Richard P Hodge, Bruce A Luxon
1Sealy Center for Structural Biology and Department of Human Biological Chemistry & Genetics, University of Texas Medical Branch at Galveston, TX 77555, USA.
Abstract:
A method using a strong anion-exchange liquid-chromatography column, Mono-Q, has been developed for high-resolution analysis and purification of oligonucleotide dithioates, which were synthesized by an automated, solid-phase, phosphorothioamidite chemistry. High-resolution separation of oligonucleotide phosphorodithioates from monothiophosphate impurities was obtained. High-resolution separation was also demonstrated at pH 8. The separation of oligonucleotide dithioates was found to be linearly dependent on the number of sulfurs for the same sequence length. Thiocyanate, SCN-, as eluting anion, can be used to purify oligonucleotides containing a high percentage of phosphorodithioate linkages in lower salt concentrations and provides better separation than chloride as eluting anion.
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