DsRNA cloning and diagnosis of beet pseudo-yellows virus by PCR and nucleic acid hybridization

Abstract

DsRNA has been extracted from beet pseudo-yellows virus infected cucumber plants, purified to homogeneity, and cDNA clones to it produced. The clones are of insufficient sensitivity to detect infection-specific RNA in dot and northern blots of crude nucleic acid extracts. However, a knowledge of the sequence of these clones has been used to synthesize oligonucleotides that have been used for polymerase chain reaction amplification of specific sequences from both purified dsRNA and from infected plants and used as a previously unavailable highly sensitive diagnostic probe. The method of cDNA synthesis has been shown to be generally applicable to some other plant viral dsRNAs and should be of use in the production of cDNA clones when virion RNA is unavailable.