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RNA interference (RNAi) is a plant self-resistant reaction and regulatory mechanism that has achieved great progress in recent years.Prokaryotic expression of dsRNA can induce RNAi, conferring the plant resistance to virus.Plants were usually infected by various viruses in their grown process.In this study, two hotspot sequences of Tobacco mosaic virus (TMV) and Potato virus Y (PVY) genomes were selected and six hpRNA vectors of single-viral or two-viral chimeric genes were constructed.The target dsRNA expressed by inducting, and then the resistant effect of the mixture dsRNA A (derived from two kinds of single-viral genes)was compared to the effect of corresponding dsRNA B (derived from one kind of two-viral chimeric gene) in different delivery ways.The data showed that dsRNA expressed by bacteria with two delivery ways could protect tobacco from co-infection with PVY and TMV, the resistant efficiency of the mixture dsRNA A was stronger than the effect of corresponding dsRNA B, the best resistant effect was 42.43%.Northern blot analysis of siRNA showed that the resistance was indeed an RNA-mediated virus resistance.