Analysis of encapsidated RNA from the Q strain of CMV (Q-CMV) has indicated the presence of a discreet population of molecules of approximately 300 nt termed RNA 5 (Peden, K. W. C., and Symons, R. H., Virology 53, 487-492, 1973.). Q-CMV RNA 5 was isolated and the 5′-end sequence was determined by direct RNA sequencing. This sequence corresponded to the exact beginning of the imperfectly conserved 3′-terminal region of genomic RNAs 1, 2, and 3. A probe generated from RNA 3 consisting of the last 130 nt of this region hybridised to RNA 5. Oligonucleotides containing sequences from the 5′-and 3′-ends of the conserved region were used to generate RNA 5 cDNA clones by RT-PCR on gel-purified RNA 5. Sequencing of these clones and primer extension analysis of transcripts (derived from the cDNA clones) and purified RNA 5 indicated that RNA 5 consists of the conserved 3′-terminal region of genomic RNAs 2 and 3.