Non-Enzymatic DNA Cleavage Reaction Induced by 5-Ethynyluracil in Methylamine Aqueous Solution and Application to DNA Concatenation

DNA can be concatenated by hybridization of DNA fragments with protruding single-stranded termini. DNA cleavage occurring at a nucleotide containing a DNA base analogue is a useful method to obtain DNA with designed protruding termini. Here, we report a novel non-enzymatic DNA cleavage reaction for DNA concatenation. We found that DNA is cleaved at a nucleotide containing 5-ethynyluracil in a methylamine aqueous solution to generate 5′-phosphorylated DNA fragment as a cleavage product. We demonstrated that the reaction can be applied to DNA concatenation of PCR-amplified DNA fragments. This novel non-enzymatic DNA cleavage reaction is a simple practical approach for DNA concatenation.

DNA可通过带有突出单链末端的DNA片段的核酸杂交实现串联拼接。在含有DNA碱基类似物的核苷酸位点发生的DNA切割反应，是获取具有预设突出末端DNA的有效方法。本文报道一种可用于DNA拼接的新型非酶促DNA切割反应。研究发现，在甲胺水溶液中，携带5-乙炔基尿嘧啶的核苷酸位点处的DNA会发生切割，切割产物为5'端磷酸化DNA片段。我们证实，该反应可应用于聚合酶链式反应（PCR）扩增所得DNA片段的拼接。该反应是一种简便实用的DNA拼接方法。