Mitochondrial minicircle DNA supports plasmid replication and maintenance in nuclei of Trypanosoma brucei.
In a search for trypanosome DNA sequences that permit replication and stable maintenance of extrachromosomal elements, a 1-kilobase-pair (kbp) fragment from a mitochondrial kinetoplast DNA (kDNA) minicircle of Trypanosoma brucei was isolated and characterized. The plasmid pTbo-1, carrying the kDNA element, is maintained in T. brucei as a supercoiled concatemer containing approximately seven to nine pTbo-1 monomer units (5.6 kbp each) in a head-to-tail orientation. The concatemer is found in approximately one copy per cell when procyclic trypanosomes are cultured in the presence of 100 micrograms of hygromycin per ml; however, in the absence of continuous hygromycin selection, the plasmid is lost from the population with a t1/2 of approximately 8.7 days (17 cell generations). A second unrelated kDNA minicircle was also able to serve as an autonomously replicating sequence (ARS) element in T. brucei, suggesting that this is a general property of kDNA minicircles. Replication of mitochondrial DNA in the nucleus may be due to either a specific consensus sequence (such as in yeast ARS elements) or nonspecific sequence characteristics (such as the degree of A&T-richness or bent DNA).
Intercampus Program in Molecular Parasitology, University of California, San Francisco 94143-1204.
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Metzenberg
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Agabian
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D017931:DNA Primers; D004261:DNA Replication; D018105:DNA, Kinetoplast / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D004272:DNA, Mitochondrial / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D016054:DNA, Protozoan / Q000302:isolation & purification; D004278:DNA, Superhelical / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism*; D015183:Restriction Mapping; D014162:Transfection; D014346:Trypanosoma brucei brucei / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D018105:DNA, Kinetoplast; D004272:DNA, Mitochondrial; D016054:DNA, Protozoan; D004278:DNA, Superhelical