申请人:Ajinomoto Co., Inc.
公开号:EP2711370A1
公开(公告)日:2014-03-26
The problem of the present invention is provision of a method of producing an n+p-mer oligonucleotide efficiently in a high yield, which includes use of, as a starting material, an n-mer oligonucleotide wherein the 3'-terminal hydroxyl group is protected, and the 5'-terminal hydroxyl group is protected by a temporary protecting group, and continuously performing, in a solution, (1) a deprotection step of the 5'-terminal hydroxyl group, (2) a 5'-terminal elongation step by the addition of a p-mer oligonucleotide wherein the 3'-position is phosphoramidited, and (3) an oxidation step or a sulfurization step of a phosphite triester moiety.
It has been found that the above-mentioned problem can be solved by adding a particular cation scavenger during the deprotection step, applying a neutralization treatment after completion of the deprotection reaction, and using a particular oxidizing agent or sulfurizing agent in the oxidation step or sulfurization step.
本发明的问题是提供一种高产率高效生产 n+p-mer 寡核苷酸的方法,该方法包括使用 n-mer 寡核苷酸作为起始材料,其中 3'末端羟基受到保护,5'末端羟基受到临时保护基团的保护、在溶液中连续进行以下步骤:(1) 5'末端羟基的去保护步骤;(2) 通过添加 p-聚合寡核苷酸进行 5'末端延伸步骤,其中 3'位被磷酸化;(3) 亚磷酸三酯分子的氧化步骤或硫化步骤。
研究发现,在脱保护步骤中加入特定的阳离子清除剂,在脱保护反应完成后进行中和处理,以及在氧化步骤或硫化步骤中使用特定的氧化剂或硫化剂,可以解决上述问题。