SELEX法筛选鼻咽癌核酸适配子的研究

目的建立鼻咽癌细胞核酸适配子库,为鼻咽癌的分子诊断、生物治疗奠定基础。方法体外合成长度为78个核苷酸的随机DNA文库,采用SELEX技术,以正常鼻咽上皮细胞为靶标进行消减筛选,以EB病毒阳性低分化鼻咽癌细胞为靶标进行10轮筛选。采用流式细胞仪检测判断核酸适配子库的特异性和亲合力,Clustal X软件分析适配子序列。结果流式细胞仪检测显示亚文库与靶细胞的结合能力随筛选轮数的增加荧光强度增强。聚类分析显示适配子可以分为3个家族,1、2家族具有保守序列。结论 EB病毒阳性鼻咽癌细胞适配子库初步建立成功,可筛选到具有较高亲和力和特异性的核酸适配子;本研究为鼻咽癌的分子诊断、生物治疗奠定了基础。

Study on selection of DNA aptamers to nasopharyngeal carcinoma cell by SELEX

Objective To establish nasopharyngeal carcinoma(NPC) nucleic acid aptamer library for laying the foundation of NPC molecular diagnosis and biological treatment.Methods An in vitro synthesized 78 mer random DNA library was subjected to subtractive selection against normal nasopharynx epithelial cell and 10 rounds of selection against NPC cells by SELEX.Binding of the aptamer pools to NPC cells was visualized by flow cytometry technique.The aptamer sequences was analyzed by clustal X.Results The subpools binding against NPC cells increased along with selection rounds.Aptamers were divided into three families by clustering analysis,and motifs were identified in 1,2 families by alignment.Conclutions We successfully established EB virus positive NPC aptamer pool,which can separate high affinity and specificity aptamers.This study lays the foundation of NPC molecular diagnosis and biological treatment.