Fig. 1.

Construction and characterization of recombinant viruses. (a) Structural organization of the MIE region of RCMV. The MIE enhancer (Enh) drives the two main IE transcripts, IE1 and IE2, by alternative splicing. (b) Isolation of mutant viruses following homologous recombination in REF/IE1 cells. (c, d) Recombinant viruses were analysed by HindIII digest, Southern blotting [using a probe directed against exon 4 (c)] and sequencing. The deletion of the IE1 protein was confirmed by Western blot analysis using a polyclonal antiserum directed against a peptide within exon 3 of the RCMV MIE region, which therefore detects both the IE1 and IE2 proteins (d). (e, f) For comparison of growth capacities, one-step [m.o.i. of 0.01 (e)] and multi-step [m.o.i. of 5 (f)] growth curves were done with WT RCMV (▴), ΔIE1ex4 (▪) and RevIE1ex4 (▴).