Isolation of Viral Double-Stranded RNAs Using a LiCl Fractionation Procedure

Abstract

A general procedure for the isolation of virus-specific double-stranded RNA (ds-RNA) is described. The procedure is based on the differential solubility of different types of nucleic acids in LiCl. Principal advantages over conventional methods are simplicity, avoidance of enzymatic treatments, and relatively good yields of undegraded ds-RNA while permitting separation of several main groups of cellular and viral nucleic acids from the same batch of tissue. The method has been successfully applied in tissues infected by several representative plant RNA viruses. The virus-specific ds-RNAs obtained have been identified by their resistance to ribo-nuclease and comparison of their electrophoretic mobilities with those of the corresponding single-stranded RNA (ss-RNA) in poly-acrylamide gels. The molecular weights of the ds-RNAs of tobacco mosaic virus, turnip yellow mosaic virus, alfalfa mosaic virus, and peanut stunt virus fit the curved log molecular weight-migration relationship constructed from a set of known marker ds-RNAs.