Development of a reverse-genetics system for Tomato spotted wilt virus

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Toward establishing an experimental system in which infection with Tomato spotted wilt virus(TSWV) can be initiated from its cDNA, we expressed TSWV nucleocapsid protein and RNA polymerase(the L protein) in Escherichia coli and Saccharomyces cerevisiae, respectively. We purified these proteins and mixed with in vitro-synthesized TSWV RNA in various conditions, however, we could not detect RNA synthesis. We also constructed plasmids to express TSWV-coded proteins and RNA in vivo, and introduced these plasmids into Nicotiana benthamiana leaves by the Agro-infiltration method, infection with TSWV did not occur. In the course of these trials, we obtained results that suggest that the translation elongation factor eEF1A plays important roles in mRNA transcription of TSWV.