以叔丁基二甲基硅烷基（TBDMS）作为5-羟甲基保护基，通过亚磷酰胺制备含有5-羟甲基-2'-脱氧胞苷修饰的DNA。
Preparation of DNA containing 5-hydroxymethyl-2'-deoxycytidine modification through phosphoramidites with TBDMS as 5-hydroxymethyl protecting group.
作者信息
Dai Qing, He Chuan
机构信息
Department of Chemistry and Institute for Biophysical Dynamics, The University of Chicago, Chicago, Illinois, USA.
出版信息
Curr Protoc Nucleic Acid Chem. 2011 Dec;Chapter 4:Unit 4.47.1-18. doi: 10.1002/0471142700.nc0447s47.
Abstract
This unit describes procedures for preparation of two phosphoramidite building blocks III and IV, both containing a TBDMS as 5-CH(2)OH-protecting group. Phosphoramidites III and IV allow efficient incorporation of 5-hmC into DNA and a "one-step" deprotection procedure to cleanly remove all the protecting groups. A "two-step" deprotection strategy is compatible with ultramild DNA synthesis, which enables the synthesis of 5 hmC-containing DNA with additional modifications. Methods are also presented for their incorporation into oligonucleotides by solid-phase synthesis, subsequent deprotection, and HPLC analysis.
摘要
本单元描述了两种亚磷酰胺砌块III和IV的制备程序，二者均含有作为5-CH(2)OH保护基团的叔丁基二甲基硅烷基（TBDMS）。亚磷酰胺III和IV可实现5-羟甲基胞嘧啶（5-hmC）高效掺入DNA，并采用“一步法”脱保护程序以彻底去除所有保护基团。“两步法”脱保护策略与超温和DNA合成兼容，能够合成带有额外修饰的含5-hmC的DNA。还介绍了通过固相合成将它们掺入寡核苷酸、随后进行脱保护以及高效液相色谱（HPLC）分析的方法。
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