REVERSE TRANSCRIPTION LOOP-MEDIATED ISOTHERMAL AMPLIFICATION (RT-LAMP) ASSAY FOR RAPID AND VISUAL DETECTION OF THE Potato Leafroll Virus
DALIA GAMIL ASEEL *
Department of Plant Protection and Biomolecular Diagnosis, Arid Lands Cultivation Research Institute (ALCRI), City of Scientific Research and Technological Applications (SRTA, City), New Borg El-Arab City, 21934, Egypt
MAHA ADEL KAWANNA
Department of Plant Pathology, Faculty of Agriculture, (EL-Shatby), Alexandria University, 21545, Alexandria, Egypt
*Author to whom correspondence should be addressed.
Abstract
Potato leafroll virus (PLRV) is a very important virus that has serious effects on the productivity of potato plants. Sensitive and efficient detection of PLRV is one of the key factors in the successful use of a viral disease control strategy. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is an innovative technique for the diagnosis of plant viruses. Here, reverse transcription-polymerase chain reaction (RT-PCR) and reverse transcription loop-mediated isothermal amplification (RT-LAMP) were used herein for the detection of PLRV in infected potato plants as well as crude plant extract. The PLRV-Coat Protein gene was detected in only two serial dilutions of cDNA (10-1 and 10-2) by RT-PCR but was detected in dilutions of up to 10−6 via RT-LAMP assay. The RT-LAMP amplified products were viewed by the naked eye, different colour agents in the infected and crude plant sample with PLRV. The specificity of the qRT-LAMP assay was also tested for Potato virus Y (PVY) and Potato virus X (PVX) infected potato plants and no reaction has occurred with PVY or PVX. The sensitivity of the qRT-LAMP (10-6) as more sensitive than the qRT-PCR (10-4) assay. The LAMP assay provides simple, efficient, low-cost, and fast detection of PLRV in infected potato plants in diagnostic laboratories or the field.
Keywords: Potato leafroll virus, RT-PCR, RT-LAMP, qRT-PCR, qRT-LAMP, betaine