寡核苷酸
PEG比率
适体
结合
核糖核酸
超滤(肾)
化学
色谱法
核苷酸
组合化学
高效液相色谱法
寡核苷酸合成
核酸
生物化学
生物
分子生物学
DNA
经济
数学分析
基因
数学
财务
作者
Stefanie Hoffmann,Johannes Hoos,Sven Klussmann,Stefan Vonhoff
标识
DOI:10.1002/0471142700.nc0446s46
摘要
This unit describes the solid-phase synthesis and downstream processing for RNA oligonucleotides with a length of up to 40 to 50 nucleotides on a 1- to 4-mmol scale with subsequent conjugation to PEG using the L-RNA spiegelmer NOX-E36 as an example. Following synthesis and two-step deprotection, the crude oligonucleotide is purified by preparative reversed-phase HPLC and desalted by tangential flow ultrafiltration. The resulting intermediate amino-modified oligonucleotide is reacted with NHS-ester-activated PEG, and the oligonucleotide-PEG conjugate is obtained after preparative AX-HPLC purification, followed by ultrafiltration and lyophilization. Critical process parameters are described, as well as time considerations and examples for analytical methods used as in-process and quality controls.
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