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Volume 13
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10.3390/bios13010111
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Article

A Designed Vessel Using Dissolvable Polyvinyl Alcohol Membrane as Automatic Valve to Couple LAMP with CRISPR/Cas12a System for Visual Detection

by
Tianyi Yang
1,
Yanju Chen
1,
Jinsong He
2,*,
Jian Wu
1,3,
Meixia Wang
4 and
Xiaoping Zhong
4,*
1
College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310058, China
2
College of Food Science and Technology, Yunnan Agricultural University, Kunming 650201, China
3
ZJU-Hangzhou Global Scientific and Technological Innovation Center, No. 733, Jianshe 3rd Road, Hangzhou 311200, China
4
Key Laboratory of Microbiol Technology and Bioinformatics of Zhejiang Province, Zhejiang Institute of Microbiology, Hangzhou 310012, China
*
Authors to whom correspondence should be addressed.
Biosensors 2023, 13(1), 111; https://doi.org/10.3390/bios13010111
Submission received: 28 November 2022 / Revised: 31 December 2022 / Accepted: 6 January 2023 / Published: 7 January 2023
(This article belongs to the Special Issue Biosensors for Monitoring Pathogenic Agents)
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Abstract

A rapid and intuitive method for detecting Vibrio parahaemolyticus (VP) was established by a designed reaction vessel which coupled CRISPR/Cas12a with loop-mediated isothermal nucleic acid amplification (LAMP). There were two spaces in the vessel-holding LAMP reaction solution and CRISPR reaction solution, respectively, which were separated with a polyvinyl alcohol (PVA) membrane. The PVA membrane could be dissolved with a water solution. The thermolabile hemolysin (TLH) gene of VP was employed as the detection target. After the target sequence of the TLH gene was amplified with LAMP, the PVA membrane would be dissolved and the CRISPR reaction solution mixed with the LAMP reaction solution. In this way, amplicons could be detected with CRISPR/Cas12a in the reaction vessel. The fluorescent signals produced by the positive samples were clearly identified by the naked eye under a UV light, while the negative samples were dark. The whole detection procedure could be finished within 35 min with a detection limit of 100 copies/µL. The designed reaction vessel is easy to produce and can effectively prevent contamination due to the opening of the reaction vessel after the LAMP reaction. Thus, it will have the potential to provide a new solution for rapid detection in the field.
Keywords: polyvinyl alcohol; dissolvable membrane; CRISPR/Cas12a; LAMP reaction; Vibrio parahaemolyticus polyvinyl alcohol; dissolvable membrane; CRISPR/Cas12a; LAMP reaction; Vibrio parahaemolyticus

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MDPI and ACS Style

Yang, T.; Chen, Y.; He, J.; Wu, J.; Wang, M.; Zhong, X. A Designed Vessel Using Dissolvable Polyvinyl Alcohol Membrane as Automatic Valve to Couple LAMP with CRISPR/Cas12a System for Visual Detection. Biosensors 2023, 13, 111. https://doi.org/10.3390/bios13010111

AMA Style

Yang T, Chen Y, He J, Wu J, Wang M, Zhong X. A Designed Vessel Using Dissolvable Polyvinyl Alcohol Membrane as Automatic Valve to Couple LAMP with CRISPR/Cas12a System for Visual Detection. Biosensors. 2023; 13(1):111. https://doi.org/10.3390/bios13010111

Chicago/Turabian Style

Yang, Tianyi, Yanju Chen, Jinsong He, Jian Wu, Meixia Wang, and Xiaoping Zhong. 2023. "A Designed Vessel Using Dissolvable Polyvinyl Alcohol Membrane as Automatic Valve to Couple LAMP with CRISPR/Cas12a System for Visual Detection" Biosensors 13, no. 1: 111. https://doi.org/10.3390/bios13010111

APA Style

Yang, T., Chen, Y., He, J., Wu, J., Wang, M., & Zhong, X. (2023). A Designed Vessel Using Dissolvable Polyvinyl Alcohol Membrane as Automatic Valve to Couple LAMP with CRISPR/Cas12a System for Visual Detection. Biosensors, 13(1), 111. https://doi.org/10.3390/bios13010111

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