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Transgenic tobacco plants have been obtained expressing nucleoprotein (N) gene sequences of three different tospoviruses known to affect vegetable crops: tomato spotted wilt virus (TSWV), tomato chlorotic spot virus (TCSV), and groundnut ringspot virus (GRSV). The chimeric plant transformation vector used comprised the three viral N gene sequences, each with a copy of the CaMV 35S promoter and the nos terminator. Despite the high levels of homology between the different N gene sequences (74-82%) and the presence of repeated promoter and terminator sequences in this construct, unrearranged copies of this triple N gene construct were stably maintained in both Escherichia coli and Agrobacterium tumefaciens plasmids used during the cloning process, as well as in several generations of transgenic tobacco plants. A transgenic tobacco line was obtained that exhibited high levels of resistance to all three tospoviruses, showing the possibility of producing transgenic plants with a broad resistance to tospoviruses by introducing tandemly cloned viral N gene sequences. DNA analysis of this transgenic plant line shows that the multivirus resistance trait is confined to a single genetic locus, which is very convenient for further breeding purposes.

Recently high levels of protection against tomato spotted wilt virus (TSWV), a negative-strand RNA virus infecting plants, have been obtained by transforming tobacco with viral nucleoprotein (N) gene sequences. Here we demonstrate that this protection is primarily due to the presence of N gene transcripts in the cells of transgenic plants, and hence appears to be RNA-mediated. Further, transgenic tobacco plants are only protected to isolates and strains of TSWV and not to other tospoviruses that share considerable nucleotide sequence homology in their N genes to TSWV. In addition to being protected after mechanical inoculation, the transgenic tobacco plants are also resistant to inoculation using viruliferous thrips, i.e. Frankliniella occidentalis (Perg.), one of the most important natural vector species.