Abstract Type I and type II DNA-topoisomerases are essential enzymes that mediate replication, transcription, recombination, and mitosis in multicellular eukaryotes but the extent of their interchange for specific reactions in vivo is controversial. Expression patterns for topoisomerase I and topoisomerase II during the embryogenesis of Drosophila melanogaster were compared with patterns of DNA replication and expression of the histone genes. In late oogenesis the maternally supplied top2 mRNA was evenly distributed throughout the egg with elevated levels at the posterior tip, a pattern that is maintained in syncytial blastoderm embryos. During gastrulation, top2 mRNA became differentially localized only to regions of DNA replication, including new expression in the gonads preceding mitosis/meiosis. Significantly higher levels of top2 mRNA were found in mitotic compared to endoreplicating tissues. The total histone mRNA was exclusively associated with DNA replication but, in contrast to top2 mRNA, mitotic and endoreplicating cells contained similar expression levels with no expression in the gonads. Striking differences exist between the distribution of the top2 mRNA and topoisomerase II protein. The protein localizes to all evolving nuclei where it persists throughout embryogenesis. A high level of top1 mRNA transcript was present without differential tissue distribution throughout embryogenesis.