DETECTION OF TOBAMOVIRUSES FROM PURIFIED OR CRUDE EXTRACTS AFTER AGAROSE-GEL ELECTROPHORESIS

Several purified tobamoviruses (U1 U2, U6, T0, CcL1) [tobacco mosaic virus and tomato mosaic virus] were separated by agarose gel electrophoresis and various staining procedures (Coomassie blue R-250, G-250, ethidium bromide, silver-based) were compared for their level of sensitivity. Coomassie blue R-250 and G-250 could detect 200 and 600 ng respectively with the common (U1) strain of tobacco mosaic virus. Ethidium bromide was less sensitive with many tobacco mosaic virus isolates, depending on various incubation factors and the nature of the virus. A simple five-step silver-based color staining of virus particles in agarose gels was developed. Our procedure was at least 20 times more sensitive than Coomassie blue R-250 and required less than 25 minutes. Coomassie blue and our silver-based staining procedures were also tested after electrophoretic separation of tobamoviruses from crude leaf extracts. The presence of 0.25 M urea in gel and buffers allowed for an efficient separation and staining of virus from tissue homogenates. Some applications are described.