Analysis of the hybridization kinetics of labeled DNA of herpes simplex virus with unlabeled excess RNA from infected cells showed that viral RNA sequences form two classes differing in molar concentration. The abundant class constituted 93.5-99.3% of total virus-specific RNA and was complementary to 14-16% of the early DNA (2 hr after infection) and to 19-22% of the late DNA (8 hr after infection) in the reproductive cycle of the virus. The early RNA sequences were found to be a subset of the late sequences. The scarce sequences constituted 0.7-6.5% of total virus-specific RNA and were complementary to 28-30% of DNA both early and late in the reproductive cycle. In this study, abundant and scarce sequences were quantitatively separated on the basis of the finding that abundant species are adenylated, i.e., contain post-transcriptionally added poly(A), whereas the scarce RNA is not. Thus, nuclear and polyribosomal adenylated RNA were complementary to 24 and 22%, respectively, of viral DNA and, in abundance competition tests, were found to compete with each other and with abundant RNA from infected cells after 8 hr. The nonadenylated polyribosomal RNA was complementary to 27% of total viral DNA of which 6% was also complementary to adenylated polyribosomal RNA. Hybridization kinetics indicated that each of the fractionated adenylated RNA formed two classes complementary to 6 and 21% of viral DNA.