A minimum length of N gene sequence in transgenic plants is required for RNA-mediated tospovirus resistance

We showed previously that transgenic plants with the green fluorescent prot ein (GFP) gene fused to segments of the nucleocapsid (N) gene of tomato spo tted wilt virus (TSWV) displayed posttranscriptional gene silencing of the GFP and N gene segments and resistance to TSWV, These results suggested tha t a chimeric transgene composed of viral gene segments might confer multipl e virus resistance in transgenic plants. To test this hypothesis and to det ermine the minimum length of the N gene that could trans-inactivate the cha llenging TSWV, transgenic plants were developed that contained GFP fused wi th N gene segments of 24-453 bp. Progeny from these plants were challenged with: (i) a chimeric tobacco mosaic virus containing the GFP gene, (ii) a c himeric tobacco mosaic virus with GFP plus the N gene of TSWV and (iii) TSW V, A number of transgenic plants expressing the transgene with GFP fused to N gene segments from 110 to 453 bp in size were resistant to these viruses . Resistant plants exhibited post-transcriptional gene silencing. in contra st, all transgenic lines with transgenes consisting of GFP fused to N gene segments of 24 or 59 bp were susceptible to TSWV, even though the transgene was post-transcriptionally silenced. Thus, virus resistance and post-trans criptional gene silencing were uncoupled when the N gene segment was 59 bp or less. These results provide evidence that multiple virus resistance is p ossible through the simple strategy of linking viral gene segments to a sil encer DNA such as GFP.