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Digestion of herpes simplex virus DNA by the HinIII or Eco RI restriction endonucleases yielded 11 to 15 fragments with molecular weights between 1 x 10(6) and 28 x10(6). The electrophoretic profiles obtained in 0.3% agarose gels with DNA fragments from none different strains of herpes simplex virus type 1 could be readily differentiated from the patterns exhibited by the corresponding fragments from four separate strains of type 2 virus; however, with each serotype, the laboratory strains differed significantly among themselves and also from isolates passaged a minimum number of times outside the human host. Digestion of all DNAs of herpes simples virus with either enzyme reproducibly generated two classes of fragments (major and minor) which differed in molar ocncentration. Moreover, although the molecular weight of an intact herpes simplex 1(F1) DNA molecule is approximately 98 x 10(6), the summed molecular weights of all major and minor HinIII fragments totalled 160 x 10(6), and the seven major fragments alone accounted for only 60 x 10(6). These unusual features indicate the existence of limited heterogeneity in the positions of cleavage sitet along individual molecules. We have eliminated the possibility that minor fragments arose from contamination with the defective DNA of high byoyant density which appears on serial undiluted passage of the virus. In fact, this latter type of DNA was resistant to cleavage by HinIII and gave large amounts of only two species of EcoRI fragments; suggesting that the defective molecules consist of many tandem repeats of a small segment of viral DNA. The heterogeneity in the viral DNA of normal density appears to be related to the structural organization of the molecules and does not necessarily imply differences in genetic content.

Anatomy of herpes simplex virus DNA: strain differences and heterogeneity in the locations of restriction endonuclease cleavage sites.