Improvement of the reverse transcription loop mediated isothermal amplification (RTLAMP) method for the detection of Peach latent mosaic viroid (PLMVd)
Boubourakas, I.N.; Fucuta, S.; Luigi, M ; Faggioli, F.; Barba, M.; Kyriakopoulou, P.E.
Proceedings of 21st International Conference on Virus and other Graft Transmissible Diseases of Fruit Crops
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julius Kuhn Archiv
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|Abstract in English:|
Peach latent mosaic viroid (PLMVd) is the most known peach viroid. Among the diagnostic techniques used for its
detection, the most recent described being the reverse transcription loop-mediated isothermal amplification method
(RT-LAMP). Several modifications were done on the basic protocol proposed by Boubourakas et al. (2009), additional
experiments were preformed in order to further evaluate the method. Namely, the reaction time was further reduced and
traces of leaf tissue, taken by a sterile toothpick, instead of tRNA were used as the intial material. Moreover, the AMV
reverse transcriptase proved to be more effective than Thermoscript, while restriction enzyme analysis was performed
on the RT-LAMP products in order to confirm that products had the respective sequences of the selected target. Finally,
the extremely high efficiency and sensitivity of RT-LAMP proved to be sufficient for the detection of PLMVd in hosts
other than peach.