Establishment and application of visual RT-LAMP technology for detecting bovine ephemeral fever virus

JIA Kun; HAN Taiguang; YUAN Liguo; SUN Lingshuang; NING Zhangyong; WANG Heng; LI Shoujun

doi:10.7671/j.issn.1001-411X.2017.03.003

Journal of South China Agricultural University > 2017 > 38(3): 15-20. > DOI: 10.7671/j.issn.1001-411X.2017.03.003

JIA Kun, HAN Taiguang, YUAN Liguo, SUN Lingshuang, NING Zhangyong, WANG Heng, LI Shoujun. Establishment and application of visual RT-LAMP technology for detecting bovine ephemeral fever virus[J]. Journal of South China Agricultural University, 2017, 38(3): 15-20. DOI: 10.7671/j.issn.1001-411X.2017.03.003

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Establishment and application of visual RT-LAMP technology for detecting bovine ephemeral fever virus

College of Veterinary Medicine, South China Agricultural University/Key Laboratory of Prevention and Control for Severe Clinical Animal Diseases of Guangdong Province/Guangdong Technological Engineering Research Center for Pet, Guangzhou 510642, China

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Received Date: June 20, 2016
Available Online: May 17, 2023

Graphical Abstract

Abstract

Abstract

Objective
To develop a rapid, convenient and visual molecular assay for detecting bovine ephemeral fever virus (BEFV).
Method
Two pairs of primers based on six conserved regions in G protein gene of BEFV were designed, and the visual reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for detecting BEFV was developed. The reaction conditions of RT-LAMP assay were optimized, and the assay was compared with PCR.
Result
The reaction ladder bands from the RT-LAMP assay were most obvious when the RT-LAMP reaction system contained 3 mmol·L^-1 Mg²⁺, 0.4 mol·L^-1 betaine and 1.2 μmol·L^-1 dNTPs mix, the ratio of inner to outer primers was 8:1, and the reaction temperature was 63 ℃. Clear reaction ladder bands were observed after 40 min of reaction. The established RT-LAMP assay had excellent specificity with only BEFV being amplified. The sensitivity was 10 times higher than that of ordinary PCR.
Conclusion
The visual RT-LAMP assay is easy to operate and highly specific, and the results can be conveniently determined. This method can be used for the rapid detection of bovine epidemic fever.
- bovine ephemeral fever,
- virus detection,
- RT-LAMP,
- PCR,
- sensitivity,
- specificity

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References (15)

References

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