Yokoe Hayato, Kokubo Kengo, Yamaoka Kazuki, Oikawa Ryota, Tomoike Fumiaki, Abe Naoko, Kimura Yasuaki, Abe Hiroshi
Department of Chemistry, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, Japan.
Research Center for Materials Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, Japan.
Commun Chem. 2025 Aug 6;8(1):232. doi: 10.1038/s42004-025-01631-x.
Chemical ligation of oligonucleotides enables assembly of long DNA constructs essential for genome engineering, DNA nanotechnology, and molecular diagnostics, but current methods often require external activators and suffer from reactive intermediate instability. Here we show a reagent-free DNA autoligation strategy based on intramolecular cross-activation between 3'-phosphorothioate (PS) and 5'-dinitrobenzenesulfonamide (DNBSA) termini on a splint DNA, yielding a P3' → N5' phosphoramidate linkage under near-physiological conditions. Ligation proceeds with over 80% yield at 37 °C and pH 8 without external reagents. The DNBSA group exhibits exceptional aqueous stability, and in situ formation of reactive intermediates contributes to high efficiency. This strategy expands the current toolkit for assembling DNA constructs and may facilitate future biotechnological and therapeutic studies.
寡核苷酸的化学连接能够组装对于基因组工程、DNA纳米技术和分子诊断至关重要的长DNA构建体，但目前的方法通常需要外部激活剂，并且存在反应中间体不稳定的问题。在此，我们展示了一种无试剂的DNA自动连接策略，该策略基于夹板DNA上3'-硫代磷酸酯（PS）和5'-二硝基苯磺酰胺（DNBSA）末端之间的分子内交叉激活，在近生理条件下产生P3'→N5'氨基磷酸酯键。在37°C和pH 8条件下，无需外部试剂，连接产率超过80%。DNBSA基团在水中具有出色的稳定性，反应中间体的原位形成有助于提高效率。该策略扩展了当前用于组装DNA构建体的工具包，并可能促进未来的生物技术和治疗研究。
