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Mareks disease virus (MDV) and reticuloendotheliosis virus (REV) can cause immune suppression in chickens and lead to great economic loss in the poultry industry.To protect chickens against both pathogens,we sought to generate recombinant Mareks disease virus type l (rMDV1) expressing the env protein of REV The complete genome of the MDV strain CVI988 was cloned as a bacterial artificial chromosome (BAC) by co-transfection with a BAC transfer vector and MDV CVI988 total DNA,and then purified by four rounds of selective passage.The infectivity of the selected BAC DNA clone was validated by MDV reconstitution in chicken embryo fibroblasts transfected with CVI988-BAC extracted from recombinant Escherichia coli.The rCVI988 had similar replication rates in vitro and pathogenicity in vivo to those of its parental strain,CVI988,whereas the rCVI988-env,which was constructed by inserting the REV env-expressing cassette into the USIO region of the CVI988 genome,replicated less than CVI988 and rCVI988.The protective efficacies ofrCVI988 and CVI988 against very virulent MDV Md5 chaUenge were similar,although the recombinant MDV-REV vaccine,rCVI988-env,has a lower protective efficacy against Md5 than the parental vaccine,it could protect a higher percentage of chickens against virulent REV challenges.Therefore,the strategy of construction of this recombinant MDV-BAC may lead to the development of recombinant vaccines against multiple viral infections.