The most abundantly transcribed HCMV gene (beta 2.7) encodes a 2.7 kb polyadenylated RNA. Although the laboratory-adapted HCMV strains AD169 and Towne possess two copies of the beta 2.7 gene within an expanded b sequence element, the low passage strain Toledo and all clinical isolates analysed contain only a single copy located within the U(L) region. A beta 2.7 deletion mutant constructed based on a strain Toledo background was shown to replicate with kinetics comparable to those of the parental virus; the beta2.7 gene is therefore not essential for virus replication in vitro. Sequencing the beta 2.7 gene from HCMV clinical isolates and the Toledo strain reveals that although the overall gene sequence is highly conserved (>99 %), the RL4 frame originally assigned in strain AD169 was disrupted in each of these viruses. Consequently, the beta 2.7 transcript does not encode any obvious translation product and thus may not function as an mRNA.