Objective In order to make a foundation for molecular diagnosis and gene engineering study of Avian Encephalomyelitis Virus(AEV),we cloned and sequenced the structural protein VP3 gene of AEV.Methods We designed a pair of primers to amplify the van-roekel VP3 gene from AEV infected SPF embryo with specific RT-PCR method.The amplified products were cloned into PDM18-T vector after purification and withdraw.Results The results showed that VP3 contains 735 bp and 245 amino acids which share 93% and 97% homology with coresponding fragments of AEV-1143 strain,respectively.We also compared it with other little RNA viruses.Conclusion Using RT-PCR method,the Van-Roekel VP3 gene in vitro has been for the first time amplified,and the VP3 gene cloned and sequenced.