The studies proposed in this application are concerned with the replication and transcription of viral DNA in cells infected with herpes simplex virus (HSV). The studies concerning viral DNA replication are designed to both characterize and map the cis functions involved in the initiation of viral DNA synthesis, and in the processing and packaging of the progeny viral DNA into mature virions. Specifically, we propose to analyze the replication and propagation of both defective and non-defective HSV DNA by using both biochemical and electron microscopic approaches. The studies concerned with viral transcription will concentrate on the analysis of the viral RNA which is derived from the S component of the HSV genome. Specifically, fine mapping of individual species of S region RNA will be performed with a combination of R looping and both standard and novel filter hybridization techniques. By comparing the structure of early and late as well as nuclear and cytoplasmic viral transcripts, we hope to answer questions concerning the maturation of S region RNA, especially with respect to splicing and to the identification of any structural signals involved in viral RNA processing. These studies will make use of selected sets of viral DNA probes derived from restricted portions of both defective and non-defective viral DNA.