Rodent malaria parasites, such as Plasmodium berghei, are practical and useful model organisms for human malaria research because of their analogies to the human malaria in terms of structure, physiology, and life cycle. Exploiting the available genetic sequence information, we constructed a cDNA library from the erythrocytic stages of P. berghei and analyzed the expressed sequence tag (EST). A total of 10,040 ESTs were generated and assembled into 2,462 clusters. These EST clusters were compared against public protein databases and 48 putative new transcripts, most of which were hypothetical proteins with unknown function, were identified. Genes encoding ribosomal or membrane proteins and purine nucleotide phosphorylases were highly abundant clusters in P. berghei. Protein domain analyses and the Gene Ontology functional categorization revealed translation/protein folding, metabolism, protein degradation, and multiple family of variant antigens to be mainly prevalent. The presently-collected ESTs and its bioinformatic analysis will be useful resources to identify for drug target and vaccine candidates and validate gene predictions of P. berghei.