Purification and electron microscopy of sowthistle yellow vein virus

Abstract

Sowthistle yellow vein virus, grown in Sonchus oleraceus L., was purified by grinding leaf material in a solution of glycine, magnesium chloride, and potassium cyanide, pH 8.1. The suspension was centrifuged 5 min at 8000 g after adjusting the pH to 8.0. Celite was added to the supernatant and the mixture was filtered through a Celite pad. The virus was then pelleted by centrifugation at high speed and after resuspending the pellets in a solution of glycine and magnesium chloride, pH 7.0, centrifuged on a density gradient. Virus was almost completely separated from contaminating plant material by density gradient electrophoresis as the final step in purification. Purified preparations contained bacilliform particles when treated with glutaraldehyde prior to negative staining with potassium phosphotungstate and uranyl acetate. When glutaraldehyde was omitted, the particles were bullet-shaped. These observations indicated that the bacilliform particle is the native form of the complete virus, whereas the bullet-shaped particle must be considered a ruptured particle. The inner component, which is apparently hollow, ends abruptly at one end; the striations at the opposite end are radially oriented. These observations suggest that the inner component has the shape of a hollow bullet. The surface projections seem to be helically arranged in two modes at the outside of the membrane. In one mode the helices make an angle of 30 ° to the long axis of the particle; in the other mode the angle is apparently 60 °. The form of the inner component may control the envelopment by a membrane.