Comovirus Isolation and RNA Extraction

Comoviruses are small, icosahedral viruses with a diameter of approx 28 nm (Fig. 1 ). Presently the genus Comovirus is comprised of 15 different viruses, of which the type species, cowpea mosaic virus (CPMV), is the most thoroughly studied (1 ,2 ). All viruses are transmitted by beetles and have a rather narrow host range. Most comoviruses have legumes as their natural hosts and usually cause mosaic or mottling symptoms. Comoviruses are mechanically transmissible, and can replicate to high levels in infected cells. Purified preparations of comoviruses consist of two, sometimes three, distinguishable particles, which can be separated by centrifugation on sucrose density gradients. These particles are designated as bottom (B), middle (M), and top (T) component, corresponding to their position in the centrifuge tube. The B- and M-component are nucleoprotein particles, each containing a segment of the single-stranded, bipartite RNA genome (denoted RNA-1 and RNA-2, respectively); T-component consists of empty protein shells. Both B- and M-components, or their RNAs, are necessary for infectivity (3 ).