Frequent homologous recombination events between molecules of one RNA component in a multipartite RNA virus

Brome mosaic bromovirus (BMV), a tripartite plus-sense RNA virus, has been used as a model system to study homologous RNA recombination among molecule s of the same RNA component. Pairs of BMV RNA3 variants carrying marker mut ations at different locations were coinoculated on a local lesion host, and the progeny RNA3 in a large number of lesions was analyzed. The majority o f doubly infected lesions accumulated the RNA3 recombinants, The distributi on of the recombinant types was relatively even, indicating that both RNA3 counterparts could serve as donor or as acceptor molecules. The frequency o f crossovers between one pair of RNA3 variants, which possessed closely loc ated markers, was similar to that of another pair of RNA3 variants with mor e distant markers, suggesting the existence of an internal recombination ho t spot, The majority of crossovers were precise, but some recombinants had minor sequence modifications, possibly marking the sites of imprecise homol ogous crossovers. Our results suggest discontinuous RNA replication, with t he replicase changing among the homologous RNA templates and generating RNA diversity. This approach can be easily extended to other RNA viruses for i dentification of homologous recombination hot spots.