摘要
目的:采用SELEX技术筛选针对人prominin-1的C-末端18肽(C18)的适配子并进行初步鉴定。方法:采用微孔板法进行SELEX筛选。用合成的KLH-C18作为筛选靶标,进行7轮筛选后进行克隆测序。选取适配子C6进行结合ELISA检测,同时进行与转染prominin-1质粒的U87细胞的免疫组化;利用链亲和素磁珠来进行适配子C6的免疫共沉淀。结果:经过7轮SELEX筛选得到具有特定二级结构的适配子。适配子C6可结合C18肽和表达人prominin-1的U87细胞。Pulldown实验证明了C6可识别天然状态的prominin-1蛋白。结论:利用"靶标切换"的SELEX技术,初步获得能与人prominin-1的C-末端18肽及其天然蛋白结合的适配子,为肿瘤的诊疗奠定实验基础。
Objective To screen and detect the aptamers against the C-terminal 18 peptides （C18） of prominin-1 by SELEX. Methods Microtiter plate method was used to screen aptamers to synthesized KLH-C18 by SELEX through 7 rounds selection and then the selected aptamers were cloned and sequenced. The C6 aptamer was selected to detect its binding to C18 peptide by ELISA method and the immunochemistry or ligand-mediated target purification by streptavidin magnetic beads were detected by C6 aptamer to the U87 cells expressing prominin-1. Results Aptamer C6 bound specifically to C 18 peptide of prominin-1 and the U87 cells expressing prominin-1. The pull clown assay verified aptamer C6 cound bind the native state of the prominin-1. Conclusion We obtained the aptamers against C-terminal 18 peptide and native state of prominin-1 by target switching method. These results could provide preliminary data for diagnosis and therapeutics of cancer.
出处
《实用医学杂志》
CAS
北大核心
2013年第4期520-522,共3页
The Journal of Practical Medicine
基金
国家自然科学基金资助项目(编号:81272509
30973481)
广东省自然科学基金资助项目(编号:07005145)
