Synthesis of short DNA and RNA fragments by resonant acoustic mixing (RAM)†

We demonstrate the first use of Resonant Acoustic Mixing (RAM) without bulk solvent for the synthesis of short oligonucleotide fragments. Using the modified H-phosphonate approach, DNA, RNA, and 2′-modified nucleotides were successfully coupled to 3′-protected nucleosides in high yields (63–92%) while reducing solvent volume by 90%. In addition to synthesizing protected phosphodiester (PO) dimers and trimers, we also synthesized protected phosphorothioate (PS) dimers in good yields (63–65%). Using phosphoramidite chemistry, we were similarly able to reduce the solvent volume by 90% while coupling DNA phosphoramidites (58–92%) and RNA phosphoramidites (55–95%) with 3′-protected nucleosides in high yields followed by traditional oxidation with iodine in solution. Both strategies were successfully scaled up to multi-gram quantities which was facilitated by the use of RAM, offering the potential for larger scale-up, up to hundreds of kilograms continuously.