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Title: Plant viroids: Isolation, Characterization/Detection, and Analysis

Author
item Owens, Robert
item SANO, TERUO - HIROSKI, JAPAN
item DURAN-VILA, NURIA - VALENCIA, SPAIN

Submitted to: Methods in Molecular Biology
Publication Type: Book / Chapter
Publication Acceptance Date: 7/1/2010
Publication Date: 1/1/2012
Citation: Owens, R.A., Sano, T., Duran-Vila, N. 2012. Plant viroids: Isolation, Characterization/Detection, and Analysis. Methods in Molecular Biology. 894:253-271.

Interpretive Summary:

Technical Abstract: When T.O. Diener discovered Potato spindle tuber viroid in 1971, only a limited number of techniques were available for plant virus detection and purification. Biological assays using indicator hosts showing strong (and characteristic) symptoms of infection and able to support high levels of viroid replication played a critical role in viroid isolation and characterization. Polyacrylamide gel electrophoresis (PAGE) was the first molecular technique to be used for the rapid (2-3 days) identification of viroid-infected plants. Because it is the only diagnostic method that is sequence-independent, PAGE under denaturing conditions continues to play a key role in the identification of new viroids. Starting in the early 1980’s, dot blot hybridization began to replace PAGE for routine viroid diagnosis. The first diagnostic protocols based on RT-PCR appeared approximately 10 years later, and much effort has subsequently been devoted to simplifying the sample preparation procedure and identifying group-specific primer pairs. This chapter describes simple, easy-to-follow protocols for each of these techniques, techniques that continue to play key roles in the discovery and characterization of new viroids.