The kinetoplast DNA of Leishmania tarentolae and Trypanosoma brucei was studied in terms of genetic organization and transcriptional activity. Several minor sequence classes of minicircles from L. tarentolae were cloned in a bacterial plasmid and were compared in terms of sequence organization. The cloned minicircles were characterized by the possession of a constant region of at least 91 nucleotides and a variable region. Minicircles from Trypanosoma brucei strain 366D were also cloned. Fragments of the maxicircle DNA from both species were also cloned in pBR322. No homology with the cloned minicircles was apparent. Several maxicircle transcripts, in addition to the 9 and 12s presumptive ribosomal RNAs, were observed in L. tarentolae. The 9 and 12s RNA genes were also mapped on the T. brucei maxicircle. Sequence homology between the L. tarentolae 9 and 12s RNA genes and the T. brucei 9 and 12s RNA genes was observed. A culture system was developed to study the developmental change of cultured bloodstream forms of T. brucei into procyclic forms. This developmental system is amenable for the study of the role of the kinetoplast DNA in the extensive mitochondrial biogenesis that occurs at this time.