Origin and direction of replication in mitochondrial DNA molecules from Drosophila melanogaster

Abstract

From a consideration of the various structural forms of partially replicated mitochondrial DNA (mtDNA) molecules from Drosophila melanogaster embryos observed in the electron microscope, it appears that the majority of molecules are replicated by a highly asymmetrical mode in which synthesis on one strand is up to 99% complete before synthesis on the second strand is initiated. Replication of the minority of molecules involves a more nearly symmetrical synthesis of the two complementary strands. The D. melanogaster mtDNA molecules have physical features with respect to which the origin and direction of replication could be mapped. These features are (i) a single region accounting for approximately 25% of the circular contour length and rich in adenine + thymine, and (ii) four EcoRI sites, all of which lie outside of this region. Molecules of this mtDNA were subjected to partial denaturation, EcoRI digestion, or partial denaturation after EcoRI digestion and the products were examined in the electron microscope. Complex forms interpretable as originating from replicative intermediates were observed. The size and structure of the components of these complex forms were wholly consistent with the interpretation that, in all of these mtDNA molecules, replication originates at, or close to, the center of the adenine + thymine-rich region and proceeds unidirectionally around the molecule toward the EcoRI site lying closest to the adenine + thymine-rich region.