oskar genes of the vector mosquitoes Anopheles gambiae, Aedes aegypti and Culex quinquefasciatus

A population replacement strategy for mosquito control requires the development of a gene drive system. The design requirements of a gene drive system are reviewed. Under investigation is the hypothesis that effective and safe gene drive systems in the form of semi-autonomous transposons can be created using the control DNA sequences of developmentally-regulated genes and currently available mosquito transformation technology. Effectiveness is defined as ability of the modified transposon to spread desired transgenes quickly into mosquito populations. Safety here refers to restriction of transposon activity to specific mosquito species, tissues and stages to mitigate environmental, ecological and social concerns regarding the release of genetically-engineered mosquitoes into the wild. The maternal effect gene oskar was identified and characterized in Anopheles gambiae, Aedes aegypti and Culex quinquefasciatus as has having potential to regulate the expression and activity of a transposable element in the mosquito germline. Techniques were developed to analyze mRNA localization patterns by hybridization in situ, and document primordial germ cell localization and migration in mosquito embryos. Mosquito oskar mRNAs localize to the posterior pole plasm of developing eggs and in the progenitor germ cells of mosquito embryos. Additionally, An. gambiae and Cx. quinquefasciatus oskar mRNAs localize transiently in the anterior of the developing embryo, a pattern not observed for D. melanogaster oskar. A 1.3 kb, 5'-end putative promoter and 0.6 kb 3'-end untranslated region isolated from Ae. aegypti oskar were used in hybrid Mos1 mariner transposon constructs to test the regulatory function of these control sequences. Subsequent analyses of transgenic lines show that the DNA control sequences used were sufficient for low levels of transgene RNA expression, but were insufficient to localize the mRNA, or activate robust protein expression. The following thesis contributes new information to mosquito embryonic development and suggests that regulatory sequences outside of the putative regulatory sequences used might be important for transcriptional and translational control of oskar genes in mosquitoes.