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United States Department of Agriculture

Agricultural Research Service

Research Project: BIOLOGICALLY-BASED TECHNOLOGIES FOR MANAGEMENT OF CROP INSECT PESTS IN LOCAL AND AREAWIDE PROGRAMS

Location: Insect Behavior and Biocontrol Research Unit

Title: A functional comparison of the 3XP3 promoter by recombinase-mediated cassette exchange in Drosophila and tephritid fly, Anastrepha suspensa

Authors
item Schetelig, Marc Florian
item Handler, Alfred

Submitted to: Genes, Genomes, and Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: February 16, 2013
Publication Date: April 1, 2013
Citation: Schetelig, M.A., Handler, A.M. 2013. A functional comparison of the 3XP3 promoter by recombinase-mediated cassette exchange in Drosophila and tephritid fly, Anastrepha suspensa. G3:Genes, Genomes, and Genomics. 3:687-693.

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 the laboratory at the USDA Agriculture Research Service, Center for Medical, Agricultural and Veterinary Entomology, Gainesville, Florida. One major problem, which contributes to the long development times for transgenic strains, is the random integration of transposon-based vectors into the genome of the insect. Such integrations often lead to gene disruptions, mutations, and ultimately to reduced fitness or loss of the strain. Development of strategies to effectively and safely utilize transgenic insects for biological control will depend upon new vector systems that can be targeted to specific genomic integration sites. This article describes the development of a new gene transfer vector system that allows the creation of target site strains to introduce site-specific target sites. Target strains will be first analyzed comprehensively and tested for host strain fitness and genomic effects on gene expression from the target site. This new vector targeting system should extend to any insect subject. This should provide a significant improvement for the development of transgenic insect strains intended for field release to control pest populations.

Technical Abstract: Transposable elements are widely used as vectors for integrating transgenes into the genome of insects. However, the random nature of transposon vector integrations often results in mutations and makes transgene expression subject to variable genomic position effects. This makes reliable quantitative comparisons of different transgenes difficult and development of highly fit transgenic strains laborious. Tools for site-specific transgene targeting are therefore essential for functional genomic comparisons and for developing the most advanced transgenic insect strains for applied use. Here we describe a recombinase-mediated cassette exchange (RMCE) gene targeting system based on CRE/loxP that is highly efficient in Drosophila, and for the first time in a non-drosophilid species, the tephritid fruit fly, Anastrepha suspensa. The system was used to compare the Drosophila constitutive polyubiquitin and the artificial 3xP3 tissue-specific promoters in the same genomic context, finding that the widely functional 3xP3 promoter is not active in the tephritid.

Last Modified: 8/27/2014
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