Recombination and linkage between structural and regulatory genes of herpes simplex virus type 1: study of the functional organization of the genome

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Abstract
Phenotypic and genetic properties of 12 markers in structural and regulatory functions of herpes simplex virus type 1 were characterized, and their recombination and segregation behavior was investigated and interpreted with reference to available information on their physical locations. The markers were ts markers in a structural glycoprotein (tsB5) and in .alpha. (immediate early; tsLB2, tsc75) or .beta. (early, delayed early; tsB1) functions with regulatory effects; together with plaque morphology (syn), phosphonoacetate resistance (Pr) and thymidine kinase (TK) phenotypes; and electrophoretically distinct variants of glycosylated (glycoprotein C, gpC; ICP10) and non-glycosylated [VP(13-14), VP23] structural and nonstructural [ICP(47-48)] polypeptides. Mean 2-factor recombination frequencies ranged from 2% (for noncomplementing mutants tsLB2 and tsc75) to 35-40% (for unlinked markers) and were influenced by the relative contributions of parental viruses to the mixed infection. Even with control of this variable, SD of mean measures of recombination frequency ranged from a minimum of 14% (with n .gtoreq. 10) to 65% (with n = 3) of mean values; no recombination freqencies higher than 55% were observed. Differences in mean 2-factor recombination frequencies between a small number of loosely linked markers were not reliable measures of real differences in linkage. Measurements of the segregation of unselected markers among recombinant progeny were used as measures of linkage. These experiments established a linkage group for markers in the long unique region of the genome additional to, but consistent with, existing physical data, i.e., TK-syn-tsB5-(tsB1.cntdot.Pr)-[gpC.cntdot.VP(13-14)]; identified markers, e.g., ICP(47-48), linked to regulatory mutations (tsLB2, tsc75) in redundant DNA sequences; and used the segregation of these regulatory mutations and linked markers among unselected progeny to demonstrate the linkage groups: Pr-syn-TK-tSc75-ICP(47-48), [VP(13-14).cntdot.gpC]-Pr-syn-TK, and TK-tsc75-[VP(13-14).cntdot.gpC]. The results were most simply explained if bi- or intermolecular recombination occurred between circular molecules or molecules catenated head-to-tail and were incompatible with intermolecular recombination as the mechanism of isomerization of herpes simplex virus DNA.