基于CRISPR-Cas12a和血糖仪的便携式生物传感器定量检测三聚氰胺  

Portable biosensor for quantitative detection of melamine based on CRISPRCas12a and a personal glucose meter

作　　者：史铠 孙国峰 周雨芹 宋九华[1] 成英[1] 尹会 贺范祥 周媛媛 李娜[1] 舒琴 陈稼轩[1] SHI Kai;SUN Guofeng;ZHOU Yuqin;SONG Jiuhua;CHENG Ying;YIN Hui;HE Fanxiang;ZHOU Yuanyuan;LI Na;SHU Qin;CHEN Jiaxuan(College of New Energy Materials and Chemistry,Leshan Normal University,Leshan 614000,China;College of Resources and Environment,Aba Teachers University,Aba 623002,China)

机构地区：[1]乐山师范学院新能源材料与化学学院,乐山614000 [2]阿坝师范学院资源与环境学院,阿坝623002

出　　处：《分析试验室》2023年第8期1088-1093,共6页Chinese Journal of Analysis Laboratory

基　　金：四川省科技创新创业苗子工程重点项目(2022JDRC0126);乐山师范学院引进人才启动项目(RC2022002);大学生创新创业训练计划项目(202210649021,202210649023,S202210649173)资助。

摘　　要：CRISPR-Cas系统作为下一代分子诊断技术在生物传感领域具有很强的应用前景。基于CRISPR-Cas12a的附属切割活性与血糖仪(PGM),构建了便携式PGM-CRISPR生物传感器,实现对三聚氰胺的定量、灵敏及特异性检测。靶标三聚氰胺与双链DNA探针中的核酸适配体特异性结合,并将另一条DNA释放出来。释放出来的DNA可以激活CRISPR-Cas12a的附属切割活性。激活后的CRISPR-Cas12a进一步切割电极上蔗糖酶修饰的单链DNA探针,将蔗糖酶修饰的DNA短片段释放到溶液中。溶液中的蔗糖酶催化蔗糖生成葡萄糖,从而产生可读取的PGM信号。在最优条件下,在0.1~2.5μmol/L范围内,PGM响应信号与三聚氰胺浓度呈良好的线性关系,检出限为37 nmol/L。该便携式PGM-CRISPR生物传感器具有很强的特异性,能够用于牛奶样品中三聚氰胺的定量检测。A CRISPR-Cas system holds great promise as a next-generation biosensing technology for molecular diagnostics.In this paper,a portable biosensor based on the trans-cleavage activity of CRISPR-Cas12a and a personal glucose meter has been developed for quantitative,sensitive and specific detection of melamine.The presence of the target melamine binds to the aptamer,leading to the release of locker DNA.And then,the leasing locker DNA activates the trans-cleavage activity of CRISPR-Cas12a to cleave the single-strand DNA(ssDNA)linker on sucrase-ssDNA modified electrode,releasing a short DNA fragment labeled with sucrase in the resulting solution.The sucrase could further catalyze sucrose to glucose,which could be detected by the PGM.Under the optimized conditions,the increase of PGM signal was relative with the concentration of melamine ranging from 0.1 to 2.5μmol/L and the limit of detection(LOD)was 37 nmol/L.Moreover,the portable biosensor has strong specificity and can be used for the quantitative detection of melamine in milk samples.

关 键 词：CRISPR-Cas12a 便携式血糖仪 三聚氰胺 

分 类 号：O657.15[理学—分析化学]