An in vitro system for the investigation of heterologous RNA recombination

Bacteriophage Phi 6 has a genome of three segments of double-stranded RNA enclosed in a polyhedral procapsid. Purified procapsids are capable of the specific packaging of viral plus strands and the synthesis of their complementary minus strands. The genomic segments of Phi 6 are capable of heterologous recombination. We have prepared an in vitro system containing purified procapsids that is capable of packaging plus strands of the genomic segments and synthesizing minus strands on these templates. The system generates heterologous recombination products when stimulated by having one of the plus strands incapable of serving as a template for minus strand synthesis. Recombinants were produced upon transfection of spheroplasts with the in vitro packaged and replicated RNA. Sites of recombination were not found to be localized in particular regions of either the donor or the recipient strands. (C) 1997 Academic Press