Infection of soybean protoplasts from cell suspension culture with bean pod mottle virus

Department of Botany and Plant Pathology Michigan State University, East Lansing, Michigan 48824, U.S.A. Protoplasts were isolated enzymically from soybean ( Glycine max L. ‘Harosoy 63’) callus continuously maintained as suspension cultures. The protoplasts were inoculated with bean pod mottle virus (BPMV) using a medium consisting of 0.4 M -sorbitol, virus, poly- L -ornithine (PLO), buffer and inorganic salts. A concentration of 0.5 x 10 5 to 1.0 x 10 5 protoplasts per ml was optimum for efficient virus infection. Although PLO was not essential for infection, it was stimulatory. A combination of PLO and CaCl 2 had what appeared to be a synergistic effect in enhancing infection, especially at low virus concentrations. When virus was preincubated for 15 min with CaCl 2 or MgCl 2 prior to inoculation, both at 0.5 m M or with potassium phosphate buffer at 10 m M pH 5.6, infection of protoplasts was significantly increased over virus which had not been preincubated. The infection process was independent of temperature under the conditions tested. BPMV and the previously introduced cowpea mosaic virus, both comoviruses, showed some contrasting differences in requirements for PLO, pH optimum and temperature. Keywords: BPMV, soybean protoplast infection Received 10 March 1981; accepted 30 July 1981.