申请人:HIRAI Kunihiro
公开号:US20120296074A1
公开(公告)日:2012-11-22
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地有效生产n+p-mer寡核苷酸的方法,该方法包括使用n-mer寡核苷酸作为起始物,其中3'-端羟基被保护,而5'-端羟基被临时保护基保护,并在溶液中连续进行以下步骤:(1) 5'-端羟基去保护步骤,(2) 通过添加一个3'-位置为磷酰胺化的p-mer寡核苷酸进行5'-端伸长步骤,和(3) 磷酸酯三酯基的氧化步骤或硫化步骤。已经发现,通过在去保护步骤中添加特定的阳离子清除剂,在去保护反应完成后施加中和处理,并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。