Construction of replication-competent Herpesvirus saimiri deletion mutants

Abstract

DNA fragments derived from the left end of Herpesvirus saimiri 11 L-DNA were cloned in Escherichia coli by using vector pBR322. Deletions were introduced within a cloned 7.4-kilobase-pair sequence by using restriction endonucleases that cut once or twice within this sequence. Permissive owl monkey kidney-cultured cells were transfected with parental strain 11 viral DNA plus cloned DNA with specific sequences deleted. By screening the progeny of these transfections with a limiting-dilution spot hybridization assay, we isolated recombinant viruses containing deletions in this region. A contiguous 4.5-kilobase-pair sequence representing 4.1% of the coding capacity of the virus was found to be unnecessary for virus replication in cultured cells. These deletion mutants will allow us to test whether sequences in this region are required for the lymphoma-inducing capacity of H. saimiri. These same procedures should also allow us to introduce foreign DNA sequences into this region for studying their expression.