Cloning and expression of Trypanosoma brucei kinetoplast DNA in Escherichia coli.

The kinetoplast DNA of Trypanosoma brucei is made of two components: mini-circles (1 kb, 90% of total kDNA) and maxi-circles (20 kb, 10%) of total kDNA). These are interlocked to form a network of about 10 000 kb. In order to analyse the components of such a network structure, we have cloned individual mini-circle molecules and two of the three EcoRI maxi-circle fragments in E. coli. Cloned mini-circles are somewhat heterogeneous in size and their restriction patterns are completely different. Despite this heterogeneity all are found to contain a homologous region(s) defined by DNA/DNA hybridization. The maxi-circles probably correspond to the mitochondrial DNA of other organisms and, in contrast to mini-circles, do not show sequence heterogeneity. One of the two cloned maxi-circle EcoRI fragments is able to direct the synthesis of two polypeptides of 10 300 and 13 500 daltons in E. coli mini-cells. Detailed analysis of this phenomenon shows that both structural genes and promoter(s) are located within the cloned maxi-circle fragment.