Development of a multiplex TaqMan probe-based real-time PCR for discrimination of variant and classical porcine epidemic diarrhea virus

Zhao, Pandeng; Bai, Juan; Jiang, Ping; Tai-shan, Tang; Li, Yufeng; Tan, Chen; Shi, Xiaoli

doi:10.1016/j.jviromet.2014.06.006

Cited by 35 publications

(29 citation statements)

References 29 publications

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“…Reverse transcription was performed using a TransScript First-Strand cDNA Synthesis Kit (Beijing TransGen Biotech Co., Ltd., China). The nano-nest PCR assay, which can distinguish between classical and variant strains of PEDV, was performed using a procedure described previously (Wang et al, 2017a). In brief, it was constructed with a nanoPCR kit (GREDBIO, China) and a mixture containing 0.4 μL cDNA, 12.5 μL nanoPCR mixture, 0.4 μL na-noTaq DNA polymerase, 1.5 μL each outer prime and nuclease-free water up to 25 μL.…”

Section: Rna Extraction Reverse Transcription and Nano-nest Pcr Detementioning

confidence: 99%

Isolation and identification of a variant subtype G 2b porcine epidemic diarrhea virus and S gene sequence characteristic

Zhu

Cao

et al. 2019

Infection, Genetics and Evolution

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Section: Rna Extraction Reverse Transcription and Nano-nest Pcr Detementioning

confidence: 99%

Isolation and identification of a variant subtype G 2b porcine epidemic diarrhea virus and S gene sequence characteristic

Zhu

Cao

et al. 2019

Infection, Genetics and Evolution

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“…An interesting concept thereby is the detection of several enteric porcine viruses, or different porcine Coronaviruses, as now exist with PEDV, TGEV, SeCoV and the Delta-Coronavirus, depending on geographical requirements Ben Salem et al, 2010). From our perspective, there seems to be little merit, however, in trying to discriminate Colorado-like and Ohio-like strains or recent and classical PEDV variants (Wang et al, 2014b;Zhao et al, 2014), as there is no difference for the control arising. Insu-lated isothermal amplification systems do not offer the ability to multiplex, but might provide the opportunity to deploy tests in the field in countries where laboratories are not easily accessible .…”

Section: Challenges For the Control Of Pedvmentioning

confidence: 99%

From the field to the lab — An European view on the global spread of PEDV

et al. 2016

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Porcine Epidemic Diarrhea Virus (PEDV) is a member of the genus Alphacoronavirus, in the family Coronaviridae, of the Nidovirales order and outbreaks of porcine epidemic diarrhoea (PED) were first recorded in England in the 1970s. Intriguingly the virus has since successfully made its way around the globe, while seemingly becoming extinct in parts of Europe before its recent return from Northern America. In this review we are re-evaluating the spread of PEDV, its biology and are looking at lessons learnt from both failure and success. While a new analysis of PEDV genomes demonstrates a wider heterogeneity of PEDV than previously anticipated with at least five rather than two genotypes, biological features of the virus and its replication also point towards credible control strategies to limit the impact of this re-emerging virus.

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“…Real-time RT-PCR with specific primer pair and probe is a more specific and sensitive technique for the identification of the presence of the virus in field samples, and also provides the capacity to quantify the virus (15). Similar assays have been developed for PEDV detection of prevalent Korean PEDV by designing primers against N gene (10) and against S gene of Chinese-prevalent strains (26). In the presented study, we established a real-time RT-PCR method that can be used to identify and quantify both classical and epidemic PEDV strains in China.…”

Section: Introductionmentioning

confidence: 99%

Development and application of a TaqMan-MGB real-time RT-PCR assay for the detection of porcine epidemic diarrhoea virus strains in China

Hou

Xie

Wang

et al. 2016

Journal of Veterinary Research

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Introduction: A real-time RT-PCR method for identification and quantification of porcine epidemic diarrhoea virus (PEDV) strains in China was developed. Material and Methods: Based on the conserved sequence of the PEDV nucleocapsid (N) gene, a primer pair and probe were designed to establish a TaqMan-MGB real-time RT-PCR assay for quantitative detection of the virus. The sequence was cloned into the pMD18-T vector and a series of diluted recombinant plasmids were used to generate a standard curve with an R2 value of 0.999. Results: The developed quantitative PCR assay detected viral titres as low as 0.1 TCID50 with high specificity and no cross-reaction with other porcine viruses (PoRV, TGEV, PRRSV, or CSFV). The intra-batch and inter-batch coefficients of variation were both less than 1%, which indicated good reproducibility. Thirty clinical diarrhoea samples obtained from pigs in Shanghai and Fujian were analysed using this quantitative PCR assay. Out of these samples, 93.3% were found to be PEDV positive. Conclusion: This approach is suitable for clinical sample identification and pathogenesis studies.

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Development of a multiplex TaqMan probe-based real-time PCR for discrimination of variant and classical porcine epidemic diarrhea virus

Cited by 35 publications

References 29 publications

Isolation and identification of a variant subtype G 2b porcine epidemic diarrhea virus and S gene sequence characteristic

Isolation and identification of a variant subtype G 2b porcine epidemic diarrhea virus and S gene sequence characteristic

From the field to the lab — An European view on the global spread of PEDV

Development and application of a TaqMan-MGB real-time RT-PCR assay for the detection of porcine epidemic diarrhoea virus strains in China

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