Abstract
Protocols for extraction and purification of viroid RNAs from the tissues of infected herbaceous plant hosts are numerous. They range from lengthy, traditional protocols that require large amounts of starting tissue and take several days to perform to those based on column chromatography which is more efficient and can be performed with smaller amounts of infected tissue. The goal of all protocols is to enrich for RNA fractions that contain viroid RNAs, and the RNA extraction procedure is chosen and adjusted for the downstream method used for detection and characterization. Removal of inhibitors/impurities is generally not an issue for herbaceous hosts unless they contain and inordinate amounts of polysaccharides, tannins, and phenols. Subsequent purification of viroid circular and linear RNAs is performed using denaturing polyacrylamide gel electrophoresis. In this chapter, a specific method routinely used for viroid purification from herbaceous hosts and problems that may be encountered is described and is intended as a reference for beginners in the field.
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References
Diener TO, Hadidi A, Owens RA (1977) Methods for studying viroids. Methods Virol 6:185–217
Niblett C, Dickson E, Horst RK, Romaine CP (1980) Additional hosts and an efficient purification procedure for four viroids. Phytopathology 70:610–615
Uyeda I, Sano T, Shikata E (1984) Purification of cucumber pale fruit viroid. Ann Phytopathol Soc Japan 50:331–338
Pallas V, Navarro A, Flores R (1987) Isolation of viroid-like RNA from hop different from hop stunt viroid. J Gen Virol 68:3210–3205
Singh RP, Kevesque D, King RR (1986) A rapid procedure for the purification of potato spindle tuber viroid—use of high performance gel permeation chromatography. Can J Plant Pathol 8:54–58
Colpan M, Schumacher J, Bruggemann W, Sanger HL, Riesner D (1983) Large-scale purification of viroid RNA using Cs2SO4 gradient centrifugation and high-performance liquid chromatography. Anal Biochem 131:257–265
Owens RA, Sano T, Duran-Vila N (2012) Plant viroids: isolation, characterization/detection, and analysis. In: Watson JM, Wang M-B (eds) Antiviral resistance in plants: methods and protocols, methods in molecular biology, vol 894, pp 252–270
Singh RP, Boucher A (1987) Electrophoretic separation of a severe from mild strains of potato spindle tuber viroid. Phytopathology 77:1588–1591
Hanold D, Vadamalai G (2017) Gel electrophoresis. In: Hadidi A, Flores R, Randles JW, Palukaitis P (eds) Viroids and satellites. Academic Press, Cambridge, pp 357–367
Navarro B, Flores R (1997) Chrysanthemum chlorotic mottle viroid: unusual structural properties of a subgroup of self-cleaving viroids with hammerhead ribozymes. Proc Natl Acad Sci U S A 94:11622–11267
Additional Reading
Palukaitis P, Symons RH (1980) Purification and characterization of the circular and linear forms of chrysanthemum stunt viroid. J Gen Virol 46:477–489
Semancik JS (1991) Chapter 22. Viroid purification and characterization. In: Graft-transmissible diseases of citrus-Handbook for detection and diagnosis of graft-transmissible diseases of citrus. FAO. www.fao.org.3.T0601E0m.htm
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Hammond, R.W. (2022). Extraction and Purification of Viroids from Herbaceous Hosts. In: Rao, A.L.N., Lavagi-Craddock, I., Vidalakis, G. (eds) Viroids. Methods in Molecular Biology, vol 2316. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1464-8_6
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DOI: https://doi.org/10.1007/978-1-0716-1464-8_6
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