Purification of Synthetic Oligodeoxynucleotides via Catching by Polymerization
Poster Mar 23, 2014
Suntara Fueangfung, Shiyue Fang
Recently, large quantities of pure synthetic oligodeoxynucleotides (ODNs) are needed for preclinical research, patient use, and clinical trials. These ODNs are synthesized on an automated synthesizer. Typically, the synthesis of ODNs generates impurities including failure sequences, which are difficult to remove. The reason is that they have the same properties as the full length ODNs. Currently, ODNs purification technologies can remove those impurities, such as reverse phase high-performance liquid chromatography (RP HPLC), anion exchange HPLC, polyacrylamide gel electrophoresis (PAGE). However, all these methods are inconvenient or costly to scale up. To solve these problems, two non-chromatographic methods of ODNs purification by polymerization are developed. In the first method, during automated synthesis, the full-length ODNs are tagged with methacrylamide group via a cleavable linker while the failure sequences are not. The full-length ODNs are incorporated into a polymer through a copolymerization process, and failure sequences and other impurities are removed by washing. The full-length ODNs are obtained by cleaving from the polymer. In the second method, the failure sequences are capped by a methacrylated phosphoramidite followed by radical acrylamide polymerization and water extraction to retrieve the full-length ODNs. The purity of ODNs has been confirmed by RP HPLC. The identity of ODNs has been established by comparing with authentic ODNs and Matrix Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) analysis. These methods do not require any expensive equipment and materials. Therefore, they are useful for large-scale drug ODN purification.
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