Development and primary application of the SYBR Green I RT-qPCR assay for the detection of the transmissible gastroenteritis virus (TGEV) S gene

Gu, Jinyang; Yue, Xiuwei; Yuan, Chen; Cui, Li; Hua, Xiuguo; Zhou, Yang; Gao, Bu-Lang; Xing, Rui

doi:10.1109/bmei.2011.6098527

Cited by 1 publication

(1 citation statement)

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“…Binding of SYBR Green and the intensity of fluorescence is proportional to the amount of amplified doublestranded DNA during the PCR amplification (21). However, a drawback is that SYBR Green can also bind to primer-dimer or non-specific amplified DNA, resulting in false positive signals (8).…”

Section: Discussionmentioning

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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Section: Discussionmentioning

confidence: 99%