|Handler, Alfred - Al|
Submitted to: Nature Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/8/2004
Publication Date: 8/8/2004
Citation: Handler, A.M., Zimowska, G.J., Horn, C. 2004. Post-integration stabilization of a transposon vector by terminal sequence deletion. Nature Biotechnology. 22:1150-1154. Interpretive Summary: The creation of transgenic strains of economically important insects for the development of more effective biological control programs is a major goal of our laboratory at the Center for Medical Agricultural and Veterinary Entomology, Gainesville, FL. Development of this methodology and strategies to effectively and safely utilize transgenic insects for biological control will depend upon new vector systems that can be stabilized to minimize strain breakdown and ecological risks related to lateral movement of the transgenes from the host species into other unintended organisms in the environment. This article describes the development of a new piggyBac gene transfer vector, that after initial integration into the genome, can have intenal sequences re-mobilized so that remaining sequences are rendered immobile. These remaining sequences will typically include genes of interest that provide the desired characteristics to the transgenic insect. This vector stabilization system is applicable to all transposon-based vectors, and should provide a significant enhancement in terms of strain stability and safety to transgenic insects intended for field release.
Technical Abstract: Transposable elements have been modified into germ-line transformation systems for nearly 20 insect species, allowing the development of transgenic insects for basic and applied studies1-3. These systems utilize a defective non-autonomous vector that results in stable vector integrations after the disappearance of transiently provided transposase helper plasmid4, which is essential to maintain true breeding lines and consistent transgene expression that would be lost after vector remobilization. This has not been a concern for laboratory studies, but the prospective use of millions of transgenic insects in biocontrol programs will likely increase the risk of remobilization by an unintended transposase source. This will be a critical concern for the ecological safety of field release programs. Here we describe an efficient method that deletes a terminal repeat sequence of a transposon vector subsequent to genomic integration. This procedure prevents transposase-mediated remobilization of the other terminal sequence and associated genes, ensuring their genomic stability.