Synthetic Errors During Solid-Phase Phosphoramidite Synthesis of Oligonucleotides

While solid-phase phosphoramidite chemistry remains the gold standard for nucleic acid synthesis, the rigorous purity requirements of modern therapeutic oligonucleotides demand a much tighter control over side reactions. This review critically examines the underlying mechanisms of impurity formation during chain assembly and evaluates practical strategies to mitigate them. We detail key nucleobase degradation pathways, specifically O⁶-guanine phosphitylation, acrylonitrile-induced alkylation, cytidine deamination, and acid-catalyzed adenine depurination. Furthermore, we address current debates surrounding the capping step, contrasting the sequence errors induced by traditional anhydride-based mixtures with the protective benefits of phosphoramidite-based capping reagents. By mapping these chemical trade-offs and evaluating various protecting group strategies, we emphasize the growing need to shift from universal synthetic cycles toward context-dependent protocols, tailoring reaction conditions to specific sequences to ensure maximum fidelity.