Programmable Aptasensor for Regulating CRISPR/Cas12a Activity

被引:12
作者
Yu, Yao [1 ]
Li, Qiaoyu [2 ,3 ]
Shi, Wen [1 ,2 ]
Yang, Yuxin [2 ]
He, Hongpeng [3 ]
Dai, Junbiao [2 ,4 ]
Mao, Guobin [2 ]
Ma, Yingxin [2 ]
机构
[1] Northwest A&F Univ, Coll Chem & Pharm, Yangling 712100, Shaanxi, Peoples R China
[2] Chinese Acad Sci, Shenzhen Inst Synthet Biol, Shenzhen Inst Adv Technol, Shenzhen Key Lab Synthet Genom,Guangdong Prov Key, Shenzhen 518055, Peoples R China
[3] Tianjin Univ Sci & Technol, Coll Biotechnol, Tianjin 300457, Peoples R China
[4] Chinese Acad Agr Sci, Agr Genom Inst Shenzhen, Minist Agr & Rural Affairs, Shenzhen Branch,Guangdong Lab Lingnan Modern Agr,K, Shenzhen 518000, Peoples R China
基金
中国国家自然科学基金;
关键词
aptamers; biosensors; CRISPR/Cas12a; molecular recognition; point-of-caretesting;
D O I
10.1021/acssensors.3c01881
中图分类号
O6 [化学];
学科分类号
0703 ;
摘要
CRISPR-mediated aptasensors have gained prevalence for detecting non-nucleic acid targets. However, there is an urgent need to develop an easily customizable design to improve the signal-to-noise ratio, enhance universality, and expand the detection range. In this article, we report a CRISPR-mediated programmable aptasensor (CPAS) platform. The platform includes single-stranded DNA comprising the aptamer sequence, locker DNA, and a crRNA recognition region, forming a hairpin structure through complementary hybridization. With T4 DNA polymerase, the crRNA recognition region was transformed into a complete double-stranded DNA through stem-loop extension, thereby activating the trans-cleavage activity of Cas 12a and generating fluorescence signals. The specific binding between the target molecule and aptamer disrupted the formation of the hairpin structure, altering the fluorescence signals. Notably, the CPAS platform allows for easy customization by simply changing the aptamer sequence and locker DNA, without entailing adjustments to the crRNA. The optimal number of bases in the locker DNA was determined to be seven nucleotides for the SARS-CoV-2 spike (S) protein and four nucleotides for ATP. The CPAS platform exhibited high sensitivity for S protein and ATP detection. Integration with a lateral flow assay enabled sensitive detection within 1 h, revealing its excellent potential for portable analysis.
引用
收藏
页码:244 / 250
页数:7
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