Development of reverse transcription loop mediated isothermal amplification assay for visual detection of Grapevine fleck virus (GFkV)

Abstract

Grapevine fleck virus (GFkV) produces a ubiquitous disease, latent in grapevine causing simple or complex infections with other more dangerous viruses. The aim of the present study is to detect GFkV through application of reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay; its efficiency has been contrasted with other procedures such as Double antibody sandwich ELISA (DAS-ELISA) and RT-polymerase chain reaction (PCR). The coat protein (CP) gene of the virus is basically used for designing the primers. Using hydroxynaphthol blue (HNB) Dye, RT-LAMP was placed in a water bath after the optimization was done. In order to detect GFkV easily and rapidly, a new immunocapture (IC)-RT-LAMP assay was developed as well; it was further compared with other assays. The results show RT-LAMP is an advantageous method because it is highly sensitive, quite cheap, user-friendly, and safe; in addition, it is performed quickly by visual detection and does not require RNA extraction (in IC-RT-LAMP).