Efficient infection from cDNA clones of cucumber mosaic cucumovirus RNAs in a new plasmid vector

Full-length cucumber mosaic cucumovirus (CMV) cDNAs were cloned into a new plasmid vector containing a modified plant virus promoter designed to transcribe the inserted sequence from its first nucleotide. cDNA copies of CMV strain Q (Q-CMV) genomic RNAs 1, 2 and 3 cloned into this vector were infectious when inoculated together, producing symptoms indistinguishable from those caused by wildtype Q-CMV infection. The infectivity of the clones could be substantially increased by excision of the viral insert together with the transcriptional promoter and terminator prior to inoculation. A diagnostic but silent mutation was introduced into the RNA 2 cDNA and found to be stably maintained in viral infection, allowing distinction of the recombinant virus from native contaminants. The infectious cDNA clones supported the replication of CMV satellite RNA when co-inoculated with biologically active Q-CMV satellite RNA transcripts. Using the infectious cDNAs described, it was found that a newly-identified overlapping gene (2b) encoded by Q-CMV RNA 2 was not essential for either systemic viral infection of Nicotiana glutinosa or replication of the satellite RNAs.