Evaluation of loop mediated isothermal amplification assay for identification of Lettuce Mosaic Virus (LMV)

Almasi, Mohammad Amin; Almasi, Galavizh

Evaluation of loop mediated isothermal amplification assay for identification of Lettuce Mosaic Virus (LMV)

Document Type : Research Paper

Authors

Young Researchers and Elites Club, North Tehran Branch, Islamic Azad University, Tehran, Iran

Abstract

The most common virus affecting lettuce (Lactuca sativa L.) in the field worldwide is Lettuce mosaic virus (LMV), belonging to the family Potyviridea, genius Potyvirus. There are several techniques to detect LMV including serological and molecular assays, but these methods take a long time and requiring sophisticated tools. The aim of this study was to reduce the time required to detect LMV, using loop-mediated isothermal amplification (LAMP) technique. Leaf samples (38 samples) with symptoms similar to LMV were collected from Kurdestan province and were subjected to a serological (DAS-ELISA) test and lastly, four samples were detected as the positive samples. Then, total RNA was extracted and one-step reverse transcription (RT)-LAMP was carried out under isothermal conditions and results were evaluated by agarose gel electrophoresis and hydroxynaphthol blue (HNB) dye. A positive result using the HNB dye was a color change in master-mix from the violet to sky blue. In addition, a novel immunocapture (IC)-RT-LAMP assay for rapid and easy detection (without RNA extraction) of LMV was developed here and its potential compared with other assays. The advantages of RT-LAMP method in compare to other methods include the high specificity, high sensitivity, high rapidity, high efficiency, safety, fascinatingly, no requirement of expensive and tools for amplification, no need to RNA extraction (in IC-RT-LAMP assay), no post-amplification treatment of the amplicons, visual detection and user friendly.

Keywords

20.1001.1.22520783.1396.7.19.5.6

Main Subjects

Plant Disease and Biotechnology

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