FIG 1.

Generation of gp150⁻ MuHV-4 expressing luciferase. (A) Two tested MuHV-4 mutations were combined. To disrupt gp150, stop codons and an EcoRI restriction site were inserted into an AfeI site at genomic position 69473 (20). To express luciferase, an M3 promoter-driven cassette was inserted into the MfeI site at genomic position 77176 (8). (B) Viral DNA was digested with EcoRI, resolved by agarose gel electrophoresis, and hybridized with ³²P-labeled probes, corresponding to nucleotides 69467 to 70918 (M7 open reading frame) of MuHV-4 and to the firefly luciferase coding sequence (pGL4.10; Promega). The open arrow shows the WT M7 fragment (13,724 bp). Black arrows show the restriction fragments containing M7-STOP. (C) Mutants were analyzed for infected-cell glycoprotein expression by flow cytometry. We used monoclonal antibodies recognizing gp150 (T1A1), gH (8C1), and gN (3F7) (14, 15, 47).