The var gene family of Plasmodium falciparum encodes the clonally variant adhesin PfEMP1 present on the surface of infected erythrocytes. A poorly understood mechanism of allelic exclusion controls the expression of PfEMP1. Transcription of var genes is developmentally and, most likely, epigenetically regulated. Here we have studied the transcriptional pattern of 28 members of this multigene family in individual parasites, early in the intraerythrocytic cycle. The results show unique patterns (type and number) of var transcripts in each individual PRBC, with 1-15 mRNA species detected per cell at 2-4 h post-invasion. When a panel of ten single PRBC was analyzed, the var gene coding for the expressed PfEMP1 was transcribed in more cells than any other, although transcripts from this gene did not give the strongest hybridization signal within each individual cell. Chromosomal mapping of transcriptionally active var genes indicated that their distribution reflects that of var loci in the genome, including a pronounced clustering in chromosome 4. These findings, taken together with existing data on var transcription at later developmental stages, suggest that the mosaic-like transcription of multiple var genes detected at the ring stage and the steady transcription of the gene encoding the expressed PfEMP1 are distinct although superimposed events, one of them random and the other taking place under some form of imprinting. With its unique features, the expression of P. falciparum var genes may reveal new principles of gene regulation.