清脆的
炸薯条
样品(材料)
联轴节(管道)
实验室晶片
微流控芯片
色谱法
生物
微流控
微生物学
副溶血性弧菌
计算机科学
化学
纳米技术
材料科学
细菌
基因
电信
生物化学
遗传学
冶金
作者
Hui Wu,Yanju Chen,Qunqing Yang,Chengbin Peng,Xiaofu Wang,Mengyao Zhang,Siwenjie Qian,Junfeng Xu,Jian Wu
标识
DOI:10.1016/j.bios.2021.113352
摘要
Vibrio parahaemolyticus ( V. parahaemolyticus ) is regarded as a major cause of seafood-associated illnesses, which has aroused widespread public concern. Here, a rapid and convenient detection method for V. parahaemolyticus detection was established by a reversible valve-assisted chip coupling with CRISPR/Cas12a. With optimized lysis buffer, loop mediated isothermal amplification (LAMP) reagents and CRISPR reagents, the whole detection process from sampling to results could be finished within 50 min. The structure of chip was simple and the cost was low. By relying on three reversible rotary valves and the rotation direction-dependent Coriolis pseudo force, the flow order of liquid and the direction of liquid flow could be precisely controlled. The LAMP amplicons were specifically and sensitively identified by CRISPR/Cas12a. Positive amplification would produce green fluorescent signal while negative amplification generated no fluorescent signal, which could be clearly distinguished by the naked eye. With 600 μL of samples processed, the limit of detection (LOD) for both pure cultured V. parahaemolyticus or spiked shrimp samples could achieve 30 copies/reaction. These illustrated the established method displayed great feasibility for real samples detection. In the future, the chip could also combine with other amplification reactions, like PCR or recombinase polymerase amplification reaction (RPA), to conduct detection by changing the corresponding lyophilized amplification reagents. Overall, the proposed detection platform displays great potential for food safety analysis and clinical diagnostics, especially in resource-limited areas. • A reversible valve-assisted chip is established for V. parahaemolyticus detection. • The structure of chip is simple and the production cost is very low. • The detection sensitivity of the proposed method can achieve 30 copies/reaction. • Amplicons are specifically and visually detected by CRISPR/Cas12a in 5 min. • The chip can be applied in different amplification reactions to conduct detection.
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