基于生物酶介导的DNA循环及滚环复制双重放大检测DNA  

Detection of DNA Based on the Enzyme Mediated Circulation and Hoop Replication Double Amplification

作　　者：牛淑妍[1] 张梦华[1] 房亚杰[1] 娄晓飞[1] NIU Shuyan ZHANG Menghua FANG Yajie LOU Xiaofei(College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, Chin)

机构地区：[1]青岛科技大学化学与分子工程学院,山东青岛266042

出　　处：《青岛科技大学学报（自然科学版）》2016年第5期479-483,共5页Journal of Qingdao University of Science and Technology:Natural Science Edition

基　　金：国家自然科学基金项目(21075073);山东省自然科学基金重点项目(ZR2010BZ006)

摘　　要：利用Nb.BbvC I限制性内切酶和Klenow聚合酶的特性设计了一个DNA的聚合剪切循环反应,再利用环DNA的滚环放大效应在金电极表面生成一条长的ssDNA。然后以氯化六氨合亚钌为电信号指示剂,设计了一个灵敏的DNA电化学生物传感器。通过循环伏安法(CV)和交流阻抗法(EIS)对电极进行表征以确保DNA的连接及杂交循环反应正常进行。在优化条件下考察了对目标DNA的响应范围及工作曲线。其检测线性范围为1.0~15nmol·L^(-1),检测限为6.62×10^(-11) mol·L^(-1)(S/N=3)。A novel and sensitive electrochemical DNA biosensor was developed for the detection of DNA based on the enzyme mediated circulation and hoop replication double amplification.The biosensor was proposed by using Hexaamimineruthenium（Ⅱ）chloride（Ru（NH3）6^2＋）as an electroactive indicator and based on the fellow two reactions：First,a cycle of DNA hybridization reaction happened by using Nb.BbvC I restriction endonuclease.Then a long ssDNA generated on gold electrode surface by using Klenow polymerase based on hoop replication.Electrodes were characterized by cyclic voltammetry（CV）and electrochemical impedance spectroscopy（EIS）to ensure that DNA was modified on the electrode surface and the hybridization reaction was carried out.The electrode response signal was detected under optimized conditions.The target DNA was quantified in a linear range from 1.0to 15nmol·L^-1,with a detection limit of 6.62×10^-11 mol·L^-1（S/N = 3）.

关 键 词：DNA电化学生物传感器 限制性内切酶 聚合酶 氯化六氨合亚钌 

分 类 号：O657.32[理学—分析化学]