Separation of potato spindle tuber viroid ribonucleic acid from Scopolia sinensis into three infectious forms and the purification and oligonucleotide pattern of fraction II RNA. Part IX

Abstract

The existence of three infectious forms of potato spindle tuber viroid (PSTV) RNA from Scopolia sinensis was demonstrated by fractionation with high salt, by reverse phase and high pressure liquid chromatography, and by polyacrylamide gel electrophoresis. Purification of fraction II was achieved by the following steps: extraction of nucleic acid with phenol, precipitation of the RNA with cetyltrimethylammonium bromide, fractionation of the RNA with lithium chloride and isopropanol, and finally gel electrophoresis. A procedure using reverse phase chromatography was developed to obtain 70–90% recovery of RNA from polyacrylamide gels. Purified PSTV fraction II RNA was digested with ribonuclease A and T1 and labelled with [γ-32P]ATP using polynucleotide kinase. The labelled digests were separated by the electrophoresis–homochromatography procedures of Sanger. About 20 and 30 spots were obtained with ribonuclease A and T1, respectively.