The Plasmodium interspersed repeat (pir) genes represent the largest multigene family in Plasmodium genomes, and the only one shared between the human pathogen, P. vivax, the simian malaria species P. knowlesi and the rodent malaria species P.y. yoelii, P. berghei and P.c. chabaudi. PIR have been shown to be expressed on the surface of red blood cells and are thought to play a role in antigenic variation. Here we have used a range of bioinformatic and experimental approaches to investigate the existence of gene subsets within P.y. yoelii pir. We have identified five groups of yir genes which could be further distinguished by chromosomal location and different alternative splicing events. Two of the groups were not highly represented among the transcribed pirs in blood stage parasites. Together these data suggest that different pir genes may be active at different stages of the life cycle of P. yoelii and may have different functions. Analysis of the 5' UTR identified a unique highly conserved yir/bir/cir specific promoter motif, which could serve as a general recognition element for yir transcription. However, its presence in front of all yirs makes it unlikely to play a role in regulating differential expression.