Development of a more efficient insect transformation system

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The long term objective of the proposed research is to develop physical and genetics systems for the delivery of cloned DNA to any insect species with subsequent site specific integration into the genome. We will explore the ability of Drosophila melanogaster embryos whose vitelline membranes have been permeabilized by a recently developed procedure to take up both DNA and nucleotide precursors. We will also utilize Drosophila embryos which are mutant for two predominant deoxyribonucleases to increase the persistence of the introduced DNA in both the somatic and germ cells. The third aim of the research is to adapt the "Flip recombinase" system from yeast for use in insect hosts. Initially Drosophila and Musca domestica (housefly) embryos will be transformed with the recombination site sequence (FRT). Then, the recombinase gene will be introduced along with selectable marker genes in plasmids containing a single FRT sequence. Site specific recombination of the marker genes will be monitored by genomic Southern and chromosomal in situ hybridization analysis. Because such a more efficient and site directed transformation system that is developed for Drosophila as a model system should be applicable to any other insect species, it will be possible to introduce genes at will into insect pests and vectors of human disease. This ability should allow biologists to devise new strategies for insect control.***//