A cytomegalovirus (CMV) isolate has been recovered from a rhesus macaque (Macaca mulatta) experimentally infected with simian immunodeficiency virus. The full-length immediate-early gene (IE) was cloned from a genomic library of this primary rhesus CMV isolate (RhCMV) using the African green monkey (AGM) CMV IE gene as probe. The promoter region and the first non-coding exon of the RhCMV IE gene have been sequenced. Nucleotide sequence comparisons from positions -733 to +625 (relative to the start site of transcription at +1) reveal 70 and 48% sequence homology with the AGM CMV IE and the human CMV (HCMV) IE genes, respectively. A region that includes the TATA box, the cis repression signal and the first exon in the RhCMV IE gene sequence is 91 and 76% homologous with the AGM CMV and HCMV IE genes, respectively. In permissive MRC-5 cells infected with RhCMV, synthesis of RNA from the IE gene peaks at 4-8 hr post infection. These studies provide a basis for elucidating molecular mechanisms regulating expression of the RhCMV IE gene and viral replication.