Isolation of replicative forms of 3' terminal subgenomic RNAs of tobacco necrosis virus

Three double-stranded RNAs of molecular weights 2.6 x 10(6), 1.05 x 10(6), and 0.94 x 10(6) are found to be synthesized during TNV infection of tobacco leaves. These double-stranded RNAs, isolated by LiCl fractionation and polyacrylamide gel electrophoresis, have been studied using RNA-RNA hybridization and RNase T(1)-resistant oligonucleotide finger-printing techniques. By these methods all three double-stranded RNAs are shown to be viral in origin and the smaller double-stranded RNA subsets of the larger RNAs. RNase T(1) oligonucleotide mapping of the viral RNA shows that both smaller double-stranded RNAs are derived from the 3' end of the viral genome. The possible role of these small double-stranded RNAs as replicative forms for mRNA synthesis and the organization and expression of the TNV genome is discussed.