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Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses
Loop-mediated isothermal amplification (LAMP) is a nucleic acid amplification technique that can be used to amplify target genes at a constant temperature, and it has several advantages, including convenience, specificity and sensitivity. However, due to the special interpretation methods of this te...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9731490/ https://www.ncbi.nlm.nih.gov/pubmed/36480573 http://dx.doi.org/10.1371/journal.pone.0278451 |
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author | Fan, Qing Xie, Zhixun Wei, You Zhang, Yanfang Xie, Zhiqin Xie, Liji Huang, Jiaoling Zeng, Tingting Wang, Sheng Luo, Sisi Li, Meng |
author_facet | Fan, Qing Xie, Zhixun Wei, You Zhang, Yanfang Xie, Zhiqin Xie, Liji Huang, Jiaoling Zeng, Tingting Wang, Sheng Luo, Sisi Li, Meng |
author_sort | Fan, Qing |
collection | PubMed |
description | Loop-mediated isothermal amplification (LAMP) is a nucleic acid amplification technique that can be used to amplify target genes at a constant temperature, and it has several advantages, including convenience, specificity and sensitivity. However, due to the special interpretation methods of this technology for reaction results, all the previously reported LAMP detection methods have been restricted to identifying a single target, which limits the application of this technology. In this study, we modified conventional LAMP to include a quencher-fluorophore composite probe complementary to the F1c segment of the inner primer FIP; upon strand separation, a gain in the visible fluorescent signal was observed. The probes could be labeled with different fluorophores, showing different colors at the corresponding wavelengths. Therefore, this multiplex LAMP (mLAMP) assay can simultaneously detect 1–3 target sequences in a single LAMP reaction tube, and the results are more accurate and intuitive. In this study, we comprehensively demonstrated a single-reaction mLAMP assay for the robust detection of three cattle viruses without nonspecific amplification of other related pathogenic cattle viruses. The detection limit of this mLAMP assay was as low as 526–2477 copies/reaction for the recombinant plasmids. It is expected that this mLAMP assay can be widely used in clinical diagnosis. |
format | Online Article Text |
id | pubmed-9731490 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-97314902022-12-09 Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses Fan, Qing Xie, Zhixun Wei, You Zhang, Yanfang Xie, Zhiqin Xie, Liji Huang, Jiaoling Zeng, Tingting Wang, Sheng Luo, Sisi Li, Meng PLoS One Research Article Loop-mediated isothermal amplification (LAMP) is a nucleic acid amplification technique that can be used to amplify target genes at a constant temperature, and it has several advantages, including convenience, specificity and sensitivity. However, due to the special interpretation methods of this technology for reaction results, all the previously reported LAMP detection methods have been restricted to identifying a single target, which limits the application of this technology. In this study, we modified conventional LAMP to include a quencher-fluorophore composite probe complementary to the F1c segment of the inner primer FIP; upon strand separation, a gain in the visible fluorescent signal was observed. The probes could be labeled with different fluorophores, showing different colors at the corresponding wavelengths. Therefore, this multiplex LAMP (mLAMP) assay can simultaneously detect 1–3 target sequences in a single LAMP reaction tube, and the results are more accurate and intuitive. In this study, we comprehensively demonstrated a single-reaction mLAMP assay for the robust detection of three cattle viruses without nonspecific amplification of other related pathogenic cattle viruses. The detection limit of this mLAMP assay was as low as 526–2477 copies/reaction for the recombinant plasmids. It is expected that this mLAMP assay can be widely used in clinical diagnosis. Public Library of Science 2022-12-08 /pmc/articles/PMC9731490/ /pubmed/36480573 http://dx.doi.org/10.1371/journal.pone.0278451 Text en © 2022 Fan et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Fan, Qing Xie, Zhixun Wei, You Zhang, Yanfang Xie, Zhiqin Xie, Liji Huang, Jiaoling Zeng, Tingting Wang, Sheng Luo, Sisi Li, Meng Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title | Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title_full | Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title_fullStr | Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title_full_unstemmed | Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title_short | Development of a visual multiplex fluorescent LAMP assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
title_sort | development of a visual multiplex fluorescent lamp assay for the detection of foot-and-mouth disease, vesicular stomatitis and bluetongue viruses |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9731490/ https://www.ncbi.nlm.nih.gov/pubmed/36480573 http://dx.doi.org/10.1371/journal.pone.0278451 |
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