SELEX技术筛选特异性结合高转移性肺癌细胞Anip973的单链DNA适配子  

In Vitro Selection of ssDNA Aptamers Against Lung Cancer Cells by SELEX

作　　者：李金明 章志怀 彭振元 

机构地区：[1]湖北省黄冈市黄州区妇女儿童医院外科,湖北黄冈438000 [2]湖北省黄冈市中心医院外科,湖北黄冈438000

出　　处：《武汉大学学报（医学版）》2011年第3期329-333,共5页Medical Journal of Wuhan University

摘　　要：目的:筛选获得一种高亲和力的能特异性结合高转移性肺癌细胞Anip973的单链DNA适配子。方法:体外合成全长88 nt的单链DNA文库,通过反复的SELEX筛选获得特异性结合Anip973细胞的单链DNA适配子,并通过生物素-辣根过氧化物酶-链霉亲和素系统初步测定筛选的适配子与细胞的亲和力,将结合力最高的适配子库进一步克隆测序。结果:经过11轮的正向筛选,以及7轮低转移性肺癌细胞AGZY83-a的反筛后,筛选获得了与Anip973细胞具有极高亲和力的适配子,亲和力从第1轮的0.08±0.02上升至0.98±0.28;将亲和力最高的第11轮适配子库进行克隆测序,将亲和力最高的适配子标记荧光素通过流式细胞术进一步验证发现筛选的ssD-NA具有很强的特异性。结论:成功筛选获得了亲和力高的特异性结合高转移性肺癌细胞Anip973的单链DNA适配子。Objective： To screen ssDNA aptamers that can specifically bind to the lung cancer cell line Anip973.Methods： A single strand DNA library with 88nt in length containing a random sequence of 30nt was synthesized in vitro.The aptamers that against the Anip973 cells were obtained by SELEX after multiple screening.The affinity between the aptamers and Anip973 cells was determined by Biotin-HRP-Streptavidin system and the ssDNA which has highest affinity was cloned and sequenced.Results： After 11 rounds positive and 7 rounds negative selection,the affinity between lung cancer cells and aptamers increased from 0.08±0.02 to 0.98±0.28.Meanwhile,the specificity between Anip973 cells and aptamers was identified by flow cytometry.Conclusion： Aptamer that bind the Anip973 cells specifically was obtained successfully.

关 键 词：SELEX技术 肺癌细胞 DNA适配子 

分 类 号：R446.6[医药卫生—诊断学]