Rapid esterification of nucleosides to solid-phase supports for oligonucleotide synthesis using uronium and phosphonium coupling reagents

Nucleosides can be esterified to solid-phase supports using uronium or phos phonium coupling reagents and a coupling additive, such as 1-hydroxybenzotr iazole (HOBT), 7-aza-1-hydroxybenzotriazole (HOAT), N-methylimidazole (NMI) , or 4-(dimethylamino)pyridine (DMAP). However, DMAP was far superior to ot her additives and high nucleoside loadings (up to 60 mu mol/g) and rapid co upling reactions (less than or equal to 10 min) were possible. Hydroxyl-der ivatized CPG was attached to nucleosides with 3'-succinyl or 3'-hydroquinon e-O O'-diacetic acid (HQDA or Q-Linker) carboxyl groups through a primary e ster linkage. Alternatively, supports derivatized with succinic acid or the Q-Linker were attached directly to the 3'-OH group of nucleosides through a secondary ester linkage. Uronium reagents (HATU or HBTU) gave the best re sults with the HQDA linker arm, while the bromophosphonium (BrOP or PyBrOP) reagents were best with the succinyl linker arm. In all cases, the couplin g reactions were much faster than previous methods using carbodiimide coupl ing reagents. The ease and speed of the reaction make this support derivati zation procedure suitable for automated in situ couplings on DNA synthesize rs.