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Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay

BACKGROUND: From April 2010 to January 2011, a severe new viral disease had devastated most duck-farming regions in China. This disease affected not only laying ducks but also meat ducks, causing huge economic losses for the poultry industry. The objective of this study is to develop a one-step reve...

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Autores principales: Wang, Youling, Yuan, Xiaoyuan, Li, Yufeng, Yu, Kexiang, Yang, Jinxing, Xu, Huaiying, Zhang, Yuxia, Yu, Kangzhen, Liao, Ming, Qin, Zhuoming
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3341576/
https://www.ncbi.nlm.nih.gov/pubmed/22185513
http://dx.doi.org/10.1186/1743-422X-8-553
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author Wang, Youling
Yuan, Xiaoyuan
Li, Yufeng
Yu, Kexiang
Yang, Jinxing
Xu, Huaiying
Zhang, Yuxia
Yu, Kangzhen
Liao, Ming
Qin, Zhuoming
author_facet Wang, Youling
Yuan, Xiaoyuan
Li, Yufeng
Yu, Kexiang
Yang, Jinxing
Xu, Huaiying
Zhang, Yuxia
Yu, Kangzhen
Liao, Ming
Qin, Zhuoming
author_sort Wang, Youling
collection PubMed
description BACKGROUND: From April 2010 to January 2011, a severe new viral disease had devastated most duck-farming regions in China. This disease affected not only laying ducks but also meat ducks, causing huge economic losses for the poultry industry. The objective of this study is to develop a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of the new virus related to Tembusu-related Flavivirus. RESULTS: The RT-LAMP assay is very simple and rapid, and the amplification can be completed within 50 min under isothermal conditions at 63°C by a set of 6 primers targeting the E gene based on the sequences analysis of the newly isolated viruses and other closely related Flavivirus.The monitoring of gene amplification can also be visualized by using SYBR green I fluorescent dye. In addition, the RT-LAMP assay for newly isolated Tembusu-related Flavivirus showed higher sensitivity with an RNA detection-limit of 2 copies/μL compared with 190 copies/μL of the conventional RT-PCR method. The specificity was identified without cross reaction to other common avian pathogens. By screening a panel of clinical samples this method was more feasible in clinical settings and there was higher positive coincidence rate than conventional RT-PCR and virus isolation. CONCLUSION: The RT-LAMP assay for newly isolated Tembusu-related Flavivirus is a valuable tool for the rapid and real-time detection not only in well-equipped laboratories but also in general conditions.
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spelling pubmed-33415762012-05-02 Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay Wang, Youling Yuan, Xiaoyuan Li, Yufeng Yu, Kexiang Yang, Jinxing Xu, Huaiying Zhang, Yuxia Yu, Kangzhen Liao, Ming Qin, Zhuoming Virol J Research BACKGROUND: From April 2010 to January 2011, a severe new viral disease had devastated most duck-farming regions in China. This disease affected not only laying ducks but also meat ducks, causing huge economic losses for the poultry industry. The objective of this study is to develop a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of the new virus related to Tembusu-related Flavivirus. RESULTS: The RT-LAMP assay is very simple and rapid, and the amplification can be completed within 50 min under isothermal conditions at 63°C by a set of 6 primers targeting the E gene based on the sequences analysis of the newly isolated viruses and other closely related Flavivirus.The monitoring of gene amplification can also be visualized by using SYBR green I fluorescent dye. In addition, the RT-LAMP assay for newly isolated Tembusu-related Flavivirus showed higher sensitivity with an RNA detection-limit of 2 copies/μL compared with 190 copies/μL of the conventional RT-PCR method. The specificity was identified without cross reaction to other common avian pathogens. By screening a panel of clinical samples this method was more feasible in clinical settings and there was higher positive coincidence rate than conventional RT-PCR and virus isolation. CONCLUSION: The RT-LAMP assay for newly isolated Tembusu-related Flavivirus is a valuable tool for the rapid and real-time detection not only in well-equipped laboratories but also in general conditions. BioMed Central 2011-12-21 /pmc/articles/PMC3341576/ /pubmed/22185513 http://dx.doi.org/10.1186/1743-422X-8-553 Text en Copyright ©2011 Wang et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research
Wang, Youling
Yuan, Xiaoyuan
Li, Yufeng
Yu, Kexiang
Yang, Jinxing
Xu, Huaiying
Zhang, Yuxia
Yu, Kangzhen
Liao, Ming
Qin, Zhuoming
Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title_full Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title_fullStr Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title_full_unstemmed Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title_short Rapid detection of newly isolated Tembusu-related Flavivirus by reverse-transcription loop-mediated isothermal amplification assay
title_sort rapid detection of newly isolated tembusu-related flavivirus by reverse-transcription loop-mediated isothermal amplification assay
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3341576/
https://www.ncbi.nlm.nih.gov/pubmed/22185513
http://dx.doi.org/10.1186/1743-422X-8-553
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