针对干扰素-γ的新型核酸适配体的筛选及鉴定

Identification of a novel DNA aptamer against IFN-γ

目的研发能特异性结合干扰素-γ(IFN-γ)的DNA适配体,为发展IFN-γ的新型检测技术提供基础。方法体外合成全长为59个碱基并含有21个随机寡核苷酸的单链DNA文库,以IFN-γ蛋白为靶标,利用指数富集的配体系统进化技术(SELEX),从单链DNA文库中筛选能够选择性结合IFN-γ蛋白的核酸适配体;流式细胞术检测富集进度、适配体与IFN-γ蛋白结合特性;MFold软件预测二级结构。结果经多轮筛选获得了能够识别IFN-γ蛋白的DNA适配体B3-8,其能够选择性地结合IFN-γ蛋白,而与血清白蛋白的结合较弱。经检测其Kd值为185 nmol/L。结论 DNA适配体B3-8能选择性地识别IFN-γ蛋白,在研发针对IFN-γ的新型检测技术方面具有应用潜能。

Objective To develop IFN-γ DNA aptamer that may potentially serve as a binding ligand in novel aptamer-based IFN-y detection. Methods A single-stranded 59nt DNA library containing 21 random oligonueleoti des was synthesized in vitro. A new aptamer B3-8 was developed by SELEX technique with IFN-γprotein as target. Flow cytometry was performed to monitor the enrichment of aptamer pool and to evaluate the binding properties of B3-8. The structure of B3-8 was predicted by MFold software. Results The aptamer B3-8 bound to the IFN-γwith a Kd of 185 nmoL/L, and had minimal cross reactivity with BSA. Conclusions B3-8 may have application poten tials in IFN-~/assay.