Rice plants showing stripe disease were collected from Ma-Anshan, Anhui province. The total RNA was extracted from samples of rice plants by TRIzol method. A pair of specific primers was designed to amplify RNA3 partial fragment of Rice stripe virus Anhui isolate by RT-PCR, and then the fragment was cloned and sequenced. Sequence analysis results showed that the fragment was consisted of 874 nts, including 636 nts of full-length gene NS3 encoding 211 amino acids. Sequence comparison showed that gene NS3 of RSV Anhui isolate shared the high sequence similarity (97.6%-99.4%) with that of RSV isolates from East China, while had relatively lower nucleotide sequence similarity (93.4%-95.4%) to that of Yunnan isolates. A phylogenetic tree based on alignment of nucleotide sequences of genes NS3 of RSV was constructed. It was found that gene NS3 of RSV Anhui isolate and those of eight RSV isolates from East China clustered into a separate branch. The results indicated that RSV isolates from East China had the closest relationship. Gene NS3 of RSV Anhui isolate was inserted into plant expression vector pBIN438, and the recombinant plant expression vector pBIN-NS3 was constructed and transformed into Agrobacterium tumefaciens. The leaves of Nicotiana benthamiana line 16c plants were co-infiltrated with A. tumefaciens mixtures carrying pBIN-NS3 and pBIN-GFP. Intense green fluorescence could be observed in the co-infiltrated patches under the UV light 6 days later. It was illustrated that the local silencing of gene GFP was inhibited, and it could be preliminarily demonstrated that protein NS3 encoded by RSV Anhui isolate was a RNA silencing suppressor.