筛选环孢霉素A适体的SELEX技术的建立(英文)

被引：8

作者：

李卫滨

兰小鹏

杨湘越

周长邵

机构：

[1] 南京军区福州总医院全军医学检验研究所

来源：

中国生物化学与分子生物学报 | 2007年 / 10期

关键词：

环孢霉素A; 适体; 随机ssDNA文库;

D O I：

10.13865/j.cnki.cjbmb.2007.10.001

中图分类号：

R91 [药物基础科学];

学科分类号：

1007 ;

摘要：

体外合成一个全长78个核苷酸,中间含35个随机序列的随机单链寡核苷酸序列(ssDNA)文库,运用指数富集的配体系统进化(SELEX)技术,以环孢霉素A(CsA)为靶目标,以磁珠作为筛选介质,利用生物素-链酶抗生物素-辣根过氧化物酶系统,检测每轮ssDNA文库与CsA的亲和力,筛选并鉴定CsA特异性的适体.经过11轮的筛选,ssDNA文库与CsA的亲和力呈上升趋势.将第10轮筛选产物克隆测序并运用相关软件进行一级结构和二级结构分析.随机挑选的19个克隆适体,根据一级结构的同源性可分为5个家族,二级结构预测以茎环(发夹)为主,这可能是适体与CsA作用的部位.CsA特异性的适体将用于酶联法、免疫荧光法等对CsA进行检测.

引用

页码：829 / 834

页数：6

共 13 条

[1]

Thrombin-binding DNA aptamer forms a unimolecular quadruplex structure in solution. Macaya RF,Schultze P,Smith FW,et al. Proceedings of the National Academy of Sciences of the United States of America . 1993

[2]

A high throughput platform for systematic evolution of ligands by exponential enrichment(SELEX). Drolet D W,Jenison R D,Smith D E,et al. Combinatorial Chemistry & High Throughput Screening . 1999

[3] ALL YOU WANTED TO KNOW ABOUT SELEX [J].

KLUG, SJ ;

FAMULOK, M .

MOLECULAR BIOLOGY REPORTS, 1994, 20 (02) :97-107

[4] FluMag-SELEX as an advantageous method for DNA aptamer selection [J].

Stoltenburg, R ;

Reinemann, C ;

Strehlitz, B .

ANALYTICAL AND BIOANALYTICAL CHEMISTRY, 2005, 383 (01) :83-91

[5]

Selection of single-stranded DNA molecules that bind and inhibit human thrombin. Bock LC,Griffin LC,Latham JA,et al. Nature . 1992

[6]

Functional requirements for specific ligand recognition by a biotin-binding RNA pseudoknot. Wilson C,Nix J,Szostak J. Biochemical Journal . 1998

[7]

An RNA motif that binds ATP. Sassanfar M,Szostak JW. Nature . 1993

[8]

RNA pseudoknots that inhibit human immunodeficiency virus type 1 reverse transcriptase. Tuerk C,Macdougal S,Gold L. Proceedings of the National Academy of Sciences of the United States of America . 1992

[9]

Systematic evolution of ligands by exponential enrichment. Tuerk C,Gold L. Science . 1990

[10]

FluMag-SELEX as an advantageous method for DNA aptamer selection. Stoltenburg R,Reinemann1 C,Strehlitz1 B. Analytical and Bioanalytical Chemistry . 2005

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