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Development of simple, rapid and sensitive detection assay for grouper nervous necrosis virus using real-time loop-mediated isothermal amplification
Mekata T, Satoh J, Inada M, Dinesh S, Harsha P, Itami T,
Published in Wiley
Volume: 38
Issue: 10
Pages: 873 - 879
A quantitative rapid detection method based on loop-mediated isothermal amplification has been developed for red-spotted grouper nervous necrosis virus (RGNNV). The nested polymerase chain reaction (PCR) assay is the mainstream inspection of the brooder in the hatchery. In this study, a real-time loop-mediated isothermal amplification (LAMP) method has been applied for RGNNV detection, known as a high-speed gene amplification procedure. Of the three temperatures (60 °C, 63 °C and 65 °C) attempted, it has been found that 63 °C is giving higher amplification from 11th minute onwards. Sensitivity analysis performed in comparison with real-time polymerase chain reaction, reverse transcriptase PCR and nested RT-PCR using various concentrations of template revealed that real-time LAMP method is efficient in terms of cost and time consumption. Specificity analysis revealed that the method developed is specific to RGNNV, whereas it has sequence cross-match with tiger puffer NNV giving advantage in detecting both the viruses. This method could be much efficient in analysing RGNNV in combination with TPNNV. © 2015 John Wiley & Sons Ltd.
About the journal
JournalData powered by TypesetJournal of Fish Diseases
PublisherData powered by TypesetWiley
Open AccessNo