The use of filter hybridization techniques for the identification, differentiation and classification of plant viruses

In attempts to use dot-blot hybridization tests for the identification of viruses or for assigning them to a certain taxonomic group we found that hybridization signals may be given not only by the homologous virus, but also by heterologous viruses belonging to the same or different taxonomic groups. Possible reasons for this phenomenon, which was observed with uncloned as well as with cloned cDNAs, are discussed. Quantitative dot-blot hybridization tests with extracted viral RNAs proved to be very sensitive in differentiating closely related viruses which were barely distinguishable in serological tests. Estimates on the degree of homology between the RNAs of different viruses may be influenced by a number of experimental parameters, such as competition for the available cDNA between homologous and heterologous RNAs or homologous RNAs in different concentrations on the same sheet of nitrocellulose, saturation phenomena due to close packaging of highly concentrated RNA on the blot and, of course, stringency conditions during washing procedures. Taking these parameters into account we have reestimated the degree of homology between the RNAs of 5 tombusviruses. Our new data suggest that the order of sequence for the relationships among these 5 tombusviruses is similar to that proposed by Koenig and Gibbs (1986) on the basis of serological data.