CRISPR/Cas12a-Mediated Rolling Circle Amplification for the Development of Liquid Crystal-Based Sensors

作者：Lijuan Zhu, Qiongzheng Hu, Zhongxing Wang, Jin‐Ming Lin, Ru‐Song Zhao · 发表于：Analytical Chemistry · 年份：2025 · DOI：10.1021/acs.analchem.5c03500 · 被引用次数：13 · 研究领域：Plant Reproductive Biology、Advanced biosensing and bioanalysis techniques、Phytoplasmas and Hemiptera pathogens

The development of high-performance liquid crystal (LC)-based sensors with remarkable sensitivity and excellent selectivity is of great importance. Herein, a CRISPR/Cas12a-based LC sensor for detecting mycotoxins in food is first reported, and the detection of aflatoxin B 1 (AFB 1 ) is chosen as a model. AFB 1 is added to magnetic beads (MBs) functionalized with double-stranded DNA (dsDNA) consisting of AFB 1 aptamer and cDNA. As the AFB 1 aptamer specifically recognizes AFB 1, cDNA is released and activates the CRISPR/Cas12a system to cut ligation DNA, thereby preventing the initiation of the rolling circle amplification (RCA) reaction. As the long-chain single-stranded DNA (ssDNA) cannot be produced to capture myristoylcholine (Myr) in the aqueous solution, a dark LC image is obtained because a Myr monolayer forms at the aqueous/LC interface. In contrast, when the RCA reaction is initiated in the absence of AFB 1, Myr in the aqueous solution is captured by long-chain ssDNA generated from the RCA reaction, causing a bright LC image. Notably, the RCA reaction on MBs exponentially amplifies the CRISPR/Cas12a-generated signals, resulting in enhanced sensitivity. The limit of detection (LOD) is about 0.312 ng/mL. Furthermore, the selectivity is greatly enhanced due to the introduction of the AFB 1 aptamer and MBs. Furthermore, a portable device is developed for rapid onsite detection. Therefore, the study provides a sensitive, selective, convenient, and promising assay for detec...