As an example of the cost-effective large-scale generation of small-interfering RNA (siRNAs), we have created transgenic tobacco plants that produce siRNAs targeted to the mRNA of the non-structural protein NS1 from the influenza A virus subtype H1N1. We have investigated if these siRNAs, specifically targeted to the 5'-portion of the NS1 transcripts (5mNS1), would suppress viral propagation in mammalian cells. Agroinfiltration of transgenic tobacco with an Agrobacterium strain harboring a 5mNS1-expressing binary vector caused a reduction in 5mNS1 transcripts in the siRNA-accumulating transgenic plants. Further, H1N1 infection of siRNA-transfected mammalian cells resulted in significant suppression of viral replication. These results demonstrate that plant-derived siRNAs can inhibit viral propagation through RNA interference and could potentially be applied in control of viral-borne diseases.