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Techniques

A Simple Purification Method for Citrus Tristeza Virus and Estimation of its Genome Size. M. Bar- Joseph, Virus Laboratory, Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel; D. J. Gumpf(2), J. A. Dodds(3), A. Rosner(4), and I. Ginzberg(5). (2)(3)Associate professors, Department of Plant Pathology, University of California, Riverside 92521; (4)Virus Laboratory, Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel; (5)Department of Neurobiology, Weizmann Institute of Science, Rehovot, Israel. Phytopathology 75:195-198. Accepted for publication 12 September 1984. Copyright 1985 The American Phytopathological Society. DOI: 10.1094/Phyto-75-195.

The citrus tristeza virus (CTV) purification procedure of Bar-Joseph et al was modified to include a short Cs₂SO₄-sucrose cushion step gradient that reduced centrifugation time and enabled rapid virus concentration and purification. Electrophoretic mobility in agarose gels of the ssRNA from purified CTV was slower than ssRNA isolated from beet yellows virus, watermelon mosaic virus, and tobacco mosaic virus. The molecular weight of CTV ssRNA was estimated to be 5.4- 6.5 x 10⁶ daltons. RNA was extracted from virus particles of various lengths and fractionated by sucrose gradient centrifugation. The RNA was dot spotted onto nitrocellulose paper and hybridized with a plasmid clone containing sequences of CTV cDNA about 600 base pairs long. The hybridization pattern of this probe did not coincide with the CTV antigen distribution as measured by ELISA but showed preferential hybridization with gradient fractions containing normal-size CTV particles.