Abstract:In order to explore the impact of the location of target sequence on RNA-mediated virus resistance, different cDNA fragments of Potato virus Y (PVY) replicase gene (nuclear inclusion b, NIb) were amplified by polymerase chain reaction (PCR), and then inserted into plant binary vector pROKII. Hairpin RNAs (hpRNAs) expression vectors were constructed. The expression vectors were introduced into Agrobacterium tumefaciens strain LBA4404 by the frozen-throwing method, and eventually introduced into tobacco NC89 via leaf disk transformation. The results of virus challenge test showed that transgenic plants transformed sequences derived from different locations of NIb gene displayed different degrees of PVY resistance; the sequences derived from the 3‘end 1/2 and the middle location mediated high levels of virus resistance, the proportion of resistant plants was above 50%; the sequences derived from the 5‘end, 5‘end 1/2 and 3‘end mediated low resistance efficiency, the proportion of resistant plants was only 10% to 30%. Northern blot showed that RNA accumulation in resistant plants was significantly lower than that in the susceptible plants, and there was a negative correlation between RNA accumulation and virus resistance, siRNAs were also detected in the resistant plants. These results indicated that the virus resistance was mediated by RNA.