Molecular genetics of herpes simplex virus: Demonstration of regions of obligatory and nonobligatory identity within diploid regions of the genome by sequence replacement and insertion

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Abstract
The DNA of herpes simplex virus (HSV) 1 and 2 consist of 2 components, L and S, each composed of unique sequences bracketed by inverted repeats. The structure of the reiterated regions of the S component was probed in marker rescue experiments involving transfection of [human and mammalian] cells with mixtures of intact HSV-1 mutant viral DNA and individual DNA fragments generated by restriction endonuclease digestion of wild-type HSV-1 or HSV-2 DNA. The results were as follows: HSV is diploid for the wild-type sequences that rescue 2 temperature-sensitive (ts) mutants. DNA fragments from both reiterated regions of the S component of HSV-1(F) DNA can rescue tsLB2 and tsD mutants. Identity of the entire reiterated sequence at both ends of S is not obligatory because only 1 end of the S component of wild phenotype virus HSV-1(1061) rescues tsD even though both ends rescue tsLB2. Genes in both reiterated sequences can be expressed. By marker rescue experiments recombinants with heterotypic ends of the S component were produced, and these specified corresponding polypeptides characteristic of both HSV-1 and HSV-2. The reiterated sequences of the S component may contain a region of obligatory identity. Thus, several recombinant clones produced by rescue with HSV-2 DNA contained identical HSV-1 DNA insertion within both reiterated regions of the HSV-1 S component. Consistent with this conclusion, the termini of the S component in the heterodiploids were identical by restriction enzyme analysis. The observation that HSV DNA can be expanded by at least 5 .times. 106 by insertion in the S component suggests that it can be a vehicle for exogenous DNA.

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