Abstract: To investigate the genomic evolution of EIAV vaccine strains during attenuation, we analyzed the sequence of env in provirus of EIAVFDDV. After amplifying env in provirus DNA of EIAVFDDV by using PCR, some positive TA clones were selected randomly to sequence and analyze. A mutation from TGG to TGA, a premature stop codon, was detected at the position of 2 128-2 130 nt of the env in 29 of 30 randomly selected clones of env. This mutation causes a truncated transmembrane protein (TM, also termed as gp45) at the 262th aa residue that resulted this mutated glycoprotein 154 residues shorter than the wild gp45. Indeed, we can see the band of predicted truncated gp45 when EIAVFDDV was analyzed by Western blot. To investigate the status of EIAV with the truncated gp45 in vivo, the env sequence of peripheral blood mononuclear cells（PBMC） associated provirus DNA and circulating virions in EIAVFDDVvaccinated horses were analyzed. TGG to TGA mutation were both found in integrated provirus DNA and the genome of the circulating virions. The above results indicate that the virion with the truncated gp45 owns the ability to infect the target cell and replicate in vivo. Extend studies are needed to understand the contribution of the truncated gp45 to immunogenicity and virulence of attenuated EIAV vaccine strains.