Representative flow chart of the strategy used to generate pBAC-BoHV-4-U. A) Through classical homologous recombination in eukaryotic cells obtained by co-transfection, the floxed (loxP sites indicated by triangles) CMV-GFP BAC cassette containing F1 plasmids elements (chloramphenicol resistant gene, Cam; F1 origin of replication, Ori2; partitioning protein genes, repE, parA, parB and parC), flanked by two homologous BoHV-4-U regions, Bo2 and Bo3, was introduced into the intergenic region of the BoHV-4-U genome [Numbers indicates sequence location in BoHV-4 genome based on the Zimmermann's sequence ]. B) Diagram of the recombinant BoHV-4-U and images of a single plaque expressing GFP (Fluorescence and phase contrast merged on fluorescence. 10×). C) Ethidium bromide stained agarose gel of three (1, 2, 3) representative HindIII digested pBAC-BoHV-4-U clones.