Characterization and cloning of the African horsesickness virus genome

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Abstract
Three mutant brome mosaic virus (BMV) RNA-2 transcripts bearing two alterations in the pseudoknot region and one in arm C of the 3′ tRNA region, previously characterized as being deficient in tRNA-like functions, have been assayed for their ability to infect and replicate (in the presence of wild-type RNAs-1 and -3) in Chenopodium hybridum plants. Although the introduced mutations have been shown to incapacitate the replication of RNA-2 in barley protoplasts, C. hybridum plants inoculated with these mutants developed local lesions indistinguishable in appearance and morphology from control inoculations containing wild-type RNA-2. Sequence analysis of progeny RNA-2 from two single lesion isolates for each mutant inoculum revealed that the input mutations were restored to functional sequences by homologous recombination within the 3′ tRNA-like region. These results, which reflect the ease with which progeny RNA can be characterized from single lesions, exemplify the value of C. hybridum for studying recombination among viral RNAs.