Structure Switchable Single Fluorophore Biosensor to Measure Dissociation Constant in PAT Aptamer Tailoring
文献类型： 外文期刊
作者： Zhu, Longjiao 1 ; Lan, Xinyue 1 ; Xiao, Xingning 3 ; Zhang, Yangzi 1 ; Du, Zaihui 1 ; Cui, Liwei 4 ; Chen, Weifeng 4 ; Wang, Meng 2 ; Huang, Kunlun 3 ; Xu, Wentao 1 ;
作者机构： 1.China Agr Univ, Dept Nutr & Hlth, Food Lab Zhongyuan, Key Lab Precis Nutr & Food Qual, Beijing 100193, Peoples R China
2.Beijing Acad Agr & Forestry Sci, Inst Qual Stand & Testing Technol, Beijing 100097, Peoples R China
3.China Agr Univ, Coll Food Sci & Nutr Engn, Key Lab Agr Genet Modified Organisms Safety Evalua, MOA, Beijing 100083, Peoples R China
4.Henan Univ Anim Husb & Econ, Sch Food & Bioengn, Zhengzhou 450046, Peoples R China
关键词：
aptamer tailoring; dissociation constant (
期刊名称：SMALL （ 影响因子：12.1； 五年影响因子：12.5 ）
ISSN： 1613-6810
年卷期： 2025 年 21 卷 30 期
页码：
收录情况： SCI
摘要： Aptamers, as synthetic oligonucleotide recognition elements, exhibit remarkable potential in biosensing applications with high specificity, chemical modifiability, and cost-effectiveness. Current approaches remain fundamentally limited by their reliance on expensive instrumentation, complex operation, and labor-intensive modification processes. Here, a structure-switchable single-fluorophore biosensor is developed integrating agarose-immobilized targets with fluorescent light-up extension primers for real-time quantitative PCR (qPCR)-based dissociation constant (Kd) determination. A 78-mer patulin aptamer is used as a model for systematic tailoring. Aptamer M1 with the best affinity (Kd = 33.41nm), obtained by removing primer regions and terminal redundant bases, exhibits a 2.5-fold increase in affinity compared to the 78 - mer. Method validation shows consistent trends, confirming reliability of the measurement platform and the efficacy of the aptamer engineering approach. Further, the molecular docking analysis identifies the central stem GC base pairs as the core interaction sites. In all, this study establishes a cost-effective, aptamer affinity quantification platform based on standard qPCR, improveing the quantitative assessment of SELEX-derived aptamers for functional element conversion, providing a robust technical framework for advancing aptamer applications in biosensing.
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