Factors Influencing the Infection of Barley Mesophyll Protoplasts with Brome Mosaic Virus RNA

Publisher Website
Google Scholar
Abstract
One of the most critical factors for the infection of barley protoplasts with brome mosaic virus (BMV) RNA was the osmotic strength of the medium during protoplast isolation and inoculation. Infection was most efficient when protoplasts were isolated and inoculated in 0.5 m-mannitol and then washed with 0.7 m-mannitol; less infection occurred when the protoplasts were isolated in 0.5 or 0.7 m-mannitol and inoculated and washed in 0.7 m-mannitol. Other conditions optimal for infection were inoculum containing 1 µg/ml BMV RNA or less, 1 µg/ml poly-l-ornithine, 20 mm-potassium citrate buffer at pH 4.7 and 0.5 mm-CaCl2, and inoculation at 0 °C. Up to 90% of protoplasts were infected in these conditions. Osmotic shock produced by increase of mannitol concentration immediately before inoculation induced invagination suggestive of pinocytosis, and this seemed to be associated with efficient infection of barley protoplasts with BMV. The discrepancy between the effect of this osmotic shock on the infection of barley protoplasts with BMV and its RNA, and the mechanism of infection are discussed.