基于石墨烯猝灭及核酸外切酶辅助循环放大适体传感器检测ATP  

Amplified Optical Aptasensor through Exonuclease Ⅲ Stimulated Regeneration of ATP Based on Graphene

作　　者：文长春[1] 赵书林[1] 黄勇[1] 潘丽[1] 陈佳[1] 胡坤[1] 

机构地区：[1]广西师范大学化学与药学学院药用资源化学与药物分子工程教育部重点实验室,广西桂林541004

出　　处：《分析测试学报》2015年第10期1168-1172,共5页Journal of Instrumental Analysis

基　　金：国家自然科学基金资助项目(21175030);药用资源化学与药物分子工程教育部重点实验室资助项目(CMEMR2014-A10)

摘　　要：基于核酸外切酶Ⅲ（ExoⅢ）辅助分析物循环放大原理,结合石墨烯（G）的超高荧光猝灭能力,以三磷酸腺苷（ATP）作为模型分析物,构建了一种新型的荧光探针,实现了ATP的高灵敏检测。两条DNA链各包含一部分ATP适体序列,其中1条DNA链的3＇端修饰有荧光染料异硫氰酸荧光素（FITC）。当模型分析物不存在时,两条DNA链通过非共价π-π堆积作用吸附在G表面,导致FITC的荧光猝灭。当目标物ATP存在时,两条DNA链与目标物ATP进行特异性结合,形成一双链夹心结构,随后加入ExoⅢ,由于ExoⅢ可对该双链夹心结构的3＇凹陷末端进行逐步水解,释放出目标物ATP和游离的FITC。释放出的目标物ATP可继续进入下一循环,不断产生游离FITC,游离FITC将脱离G表面,从而导致体系的荧光恢复,并且恢复的荧光强度与ATP浓度在0.02~1μmol/L范围内存在良好的线性关系,检出限（S/N=3）为9 nmol/L。A highly sensitive platform for adenosine triphosphate（ ATP） assay was designed based on exonuclease Ⅲ（ Exo Ⅲ） induced target recycling amplification and fluorescence quenching of graphene. The system mainly consists of two nucleic acid strands P1 and P2carrying a part of aptamer sequences for ATP,and P1 with FITC labeled on its 3＇ end. Upon addition of target ATP to the system,P1 hybridizes with P2,and forms a DNA duplex sandwich structure. The formation of DNA duplex sandwich structure triggers selective enzymatic cleavage of 3＇- recessed end by Exo Ⅲ,resulting in the release of target ATP and FITC. Released target ATP then hybridizes with nucleic acid strands P1 and P2,generating many FITC molecule. As a result,the FITC molecule break away from the G surface,and lead to the fluorescence recovery of the dye FITC. This assay exhibits a high sensitivity with a detection range of 0. 02- 1 μmol / L and a detection limit（ S / N = 3） of 9 nmol / L.

关 键 词：三磷酸腺苷 核酸外切酶Ⅲ 石墨烯 荧光法 适体传感器 

分 类 号：O657.3[理学—分析化学] Q525[理学—化学]