Author
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SCHETELIG, MARC - Former ARS Employee |
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Handler, Alfred - Al |
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Submitted to: Genetica
Publication Type: Peer Reviewed Journal Publication Acceptance Date: 5/20/2013 Publication Date: 6/3/2013 Citation: Schetelig, M., Handler, A.M. 2013. Germline transformation of the spotted wing drosophilid, Drosophila suzukii, with a piggyBac transposon vector. Genetica. 141(4-6):189-193. Interpretive Summary: The ability to achieve gene transfer in economically important insects is a major goal of the USDA-Agriculture Research Service, Center for Medical, Veterinary and Agricultural Entomology in Gainesville, Florida. A biocontrol strategy for the population suppression of pest flies is the sterile insect technique (SIT). The primary needs for successful SIT is the ability to mass rear and release large numbers of sterile, though sexually active, adult male insects, and to eliminate females early in development to avoid rearing costs and their subsequent sterilization and release. The ability to genetically manipulate fruit flies allows the creation of transgenic conditional lethal strains that eliminate females during rearing, and sterilize surviving males. A first step in development of SIT for spotted-wing Drosophila, an emerging pest of small fruit, is a piggyBac transformation using a vector carrying a female-specific lethal gene and green and blue fluorescent protein gene markers for GMO detection. Transformation was achieved at a frequency of ~16%, with transformants easily detectable. It is expected that this transformed strain will be used in a female-specific lethal SIT strategy for spotted wing Drosophila. Technical Abstract: Drosophila suzukii is a pest of small fruits in many parts of the world, whose management is limited to cultural practices and the use of insecticides. Here we describe a method to genetically manipulate this species in the first step to create female lethality strains useful for the sterile insect technique (SIT) method of population suppression. This was achieved by the germ-line transformation of D. suzukii with a piggyBac transposon vector having a female-specific lethality effector construct. This can be used in a tetracycline-suppressible conditional gene expression system, when crossed to a suitable tet-transactivator strain. Transformation occurred efficiently, at a frequency of 16% per fertile G0 embryo injected with vector and helper transposase plasmids. The vector was marked for transformant selection with the polyubiquitin regulated EGFP fluorescent protein, and contains the attP landing site and heterospecific lox recombination sites for post-integration modification of the transgene vector. The 3xP3-AmCyan fluorescent protein marker was inserted within the lox sites to follow a possible recombinase-mediated cassettte exchange, that would allow subsequent improvement of the transgenic strain by immobilization of the vector and introduction of new marker cassettes. |
