Herpes simplex virus (HSV) type 2 (strain HG52) has four I sites at map coordinates (m.c.) 0.45, 0.7, 0.91 and 0.94, i.e. two in the unique long and two in the unique short regions of the genome. Previously, we had isolated a genome containing only the 0.45 and 0.94 I sites. Here we report the isolation of a mutant (JH2611) in which all four I sites have been removed using an enrichment selection procedure, without any loss of viability. Removal of each site has been shown to be due to a base change or small undetectable deletion/insertion of less than 100 bp. In HSV-1, the I site at 0.45 m.c. is in an open reading frame coding for a polypeptide of 14K. Both the 0.7 and 0.94 m.c. HSV-2 I sites are in intergenic positions. The 0.91 m.c. I site has been shown to be within the coding sequence of the glycoprotein gG-2. Synthesis of gG-2 by JH2611 and two other mutants, JH2610 (formerly HG52X163X3) and JH2609 (formerly HG52X163X21), in which the 0.91 m.c. site has been deleted was analysed by immunoprecipitation using the gG-2-specific monoclonal antibodies AP1 and LP5 and the anti-peptide serum 14713. In the mutants JH2610 and JH2611 neither gG-2 nor its precursor were detected but the monoclonal antibodies detected two polypeptides migrating above the normal positions of gG-2 and the gG-2 precursor; these were not precipitated by the anti-peptide serum. With the mutant JH2609 neither gG-2 nor its precursors could be detected by either the monoclonal antibodies or the anti-peptide serum. The results strongly suggest that gG-2 is non-essential for the growth of HSV-2 .


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