A New Sensitive Method for Detection of Viroids

Background and Aims: Viroids are smallest known plant pathogens and cause several economically significant diseases. Until recently, viroid detection relied mainly on biological tests and indexing. Today various diagnostic techniques such as nucleic acid hybridization, southern blot and reverse transcription coupled with polymerase chain reaction (RT-PCR) are being used for detection and diagnosis of viroids. Methods: This paper describes a new method for detection of citrus viroids, based on a combination of RT-PCR and dot blot hybridization (RT-PCR-DBH). In this method instead of using nucleic acid extracted directly from the plants, RT-PCR products are subjected to dot-blot hybridization. Results: The results showed that the above mentioned method has some advantages compared with the other methods. It is more sensitive, relatively simple, cost-effective, rapid and easy to apply. It was about 1000 times more sensitive than southern blot and about 100 times more sensitive than PCR in detecting hop stunt viroid in citrus. Conclusion: The introduced method here has a high potential in diagnosis of viroids and is suitable for detection of low concentrations of the agent.