Rapid and sensitive detection of porcine epidemic diarrhea virus by reverse transcription loop-mediated isothermal amplification combined with a vertical flow visualization strip
| Autor: | Jinding Chen, Mingqiu Zhao, Jiaying Wang, Hongchao Gou, Deng Jieru, Jingjing Pei, Wenjun Liu |
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| Rok vydání: | 2015 |
| Předmět: |
Swine
Loop-mediated isothermal amplification Sensitivity and Specificity chemistry.chemical_compound Animals Molecular Biology Pathogen Reverse Transcription Loop-mediated Isothermal Amplification Swine Diseases Detection limit biology Porcine epidemic diarrhea virus Reverse Transcription Cell Biology Nucleic acid amplification technique biology.organism_classification Virology Molecular biology Reverse transcriptase Calcein Molecular Diagnostic Techniques chemistry RNA Viral Coronavirus Infections Nucleic Acid Amplification Techniques |
| Zdroj: | Molecular and Cellular Probes. 29:48-53 |
| ISSN: | 0890-8508 |
| DOI: | 10.1016/j.mcp.2014.11.004 |
| Popis: | Porcine epidemic diarrhea virus (PEDV) is an important pathogen that causes vomiting, diarrhea, and dehydration, leading to serious damage to the swine industry worldwide. The establishment of effective diagnostic methods is imperative. However, traditional methods are often unsuitable. In this study, reverse transcription loop-mediated isothermal amplification (RT-LAMP) was combined with a vertical flow (VF) nucleic acid detection strip to detect PEDV. Parameters that affect the RT-LAMP reaction were optimized. The RT-LAMP-VF assay that we established was performed at 62 °C for 40 min, and then directly evaluated on the VF visualization strip cassette. The method demonstrated high specificity for PEDV. The detection limit was 10 pg of ribonucleic acid, consistent with RT-PCR, RT-LAMP detected products on agarose gels and by direct calcein fluorescence. Application of this method to clinical samples yielded a positivity rate that was comparable to that obtained for RT-PCR. This technique saves time and is efficient, and is thus expected to be useful for the diagnosis of PEDV infection in the field. |
| Databáze: | OpenAIRE |
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