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 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 cassette exchange, that would allow subsequent improvement of the transgenic strain by immobilization of the vector and introduction of new marker cassettes.
References
Berghammer AJ, Klingler M, Wimmer EA (1999) A universal marker for transgenic insects. Nature 402:370–371
Goodhue RE, Bolda M, Farnsworth D, Williams JC, Zalom FG (2011) Spotted wing drosophila infestation of California strawberries and raspberries: economic analysis of potential revenue losses and control costs. Pest Manag Sci 67:1396–1402
Groth AC, Fish M, Nusse R, Calos MP (2004) Construction of transgenic Drosophila by using the site-specific integrase from phage phiC31. Genetics 166:1775–1782
Handler AM (2000) An introduction to the history and methodology of insect gene transfer. In: Handler AM, James AA (eds) Insect transgenesis: methods and applications. CRC Press LLC, Boca Raton, pp 3–26
Handler AM, Harrell RA (1999) Germline transformation of Drosophila melanogaster with the piggyBac transposon vector. Insect Mol Biol 8:449–457
Handler AM, Harrell RA (2001a) Polyubiquitin-regulated DsRed marker for transgenic insects. Biotechniques 31(820):824–828
Handler AM, Harrell RA (2001b) Transformation of the Caribbean fruit fly, Anastrepha suspensa, with a piggyBac vector marked with polyubiquitin-regulated GFP. Insect Biochem Mol Biol 31:199–205
Horn C, Wimmer EA (2003) A transgene-based, embryo-specific lethality system for insect pest management. Nat Biotechnol 21:64–70
Knipling EF (1955) Possibilities of insect control or eradication through the use of sexually sterile males. J Econ Entomol 48:459–462
Knipling EF (1959) Sterile-male method of population control. Science 130:902–904
Krafsur ES (1998) Sterile insect technique for suppressing and eradicating insect population: 55 years and counting. J Agric Entomol 15:303–317
Lee JC, Bruck DJ, Dreves AJ, Ioriatti C, Vogt H, Baufeld P (2011) In focus: spotted wing drosophila, Drosophila suzukii, across perspectives. Pest Manag Sci 67:1349–1351
Lorenzen MD, Berghammer AJ, Brown SJ, Denell RE, Klingler M, Beeman RW (2003) piggyBac-mediated germline transformation in the beetle Tribolium castaneum. Insect Mol Biol 12:433–440
O’Brochta DA, Pilitt KL, Harrell RA, 2nd, Aluvihare C, Alford RT (2012) Gal4-based enhancer-trapping in the malaria mosquito Anopheles stephensi. G3 (Bethesda) 2:1305–1315
Oberstein A, Pare A, Kaplan L, Small S (2005) Site-specific transgenesis by Cre-mediated recombination in Drosophila. Nat Methods 2:583–585
O’Brochta DA, Alford RT, Pilitt KL, Aluvihare CU, Harrell RA 2nd (2011) piggyBac transposon remobilization and enhancer detection in Anopheles mosquitoes. Proc Natl Acad Sci USA 108:16339–16344
Ogaugwu CE, Schetelig MF, Wimmer EA (2013) Transgenic sexing system for Ceratitis capitata (Diptera: Tephritidae) based on female-specific embryonic lethality. Insect Biochem Mol Biol 43:1–8
Rubin GM, Spradling AC (1982) Genetic transformation of Drosophila with transposable element vectors. Science 218:348–353
Schetelig MF, Handler AM (2012a) Strategy for enhanced transgenic strain development for embryonic conditional lethality in Anastrepha suspensa. Proc Natl Acad Sci USA 109:9348–9353
Schetelig MF, Handler AM (2012b) A transgenic embryonic sexing system for Anastrepha suspensa (Diptera: Tephritidae). Insect Biochem Mol Biol 42:790–795
Schetelig MF, Handler AM (2013) A functional comparison of the 3xP3 promoter by recombinase-mediated cassette exchange in Drosophila and a tephritid fly, Anastrepha suspensa. G3. doi:10.1534/g3.112.005488
Schetelig MF, Caceres C, Zacharopoulou A, Franz G, Wimmer EA (2009a) Conditional embryonic lethality to improve the sterile insect technique in Ceratitis capitata (Diptera: Tephritidae). BMC Biol 7:4
Schetelig MF, Scolari F, Handler AM, Kittelmann S, Gasperi G, Wimmer EA (2009b) Site-specific recombination for the modification of transgenic strains of the Mediterranean fruit fly Ceratitis capitata. Proc Natl Acad Sci USA 106:18171–18176
Scolari F, Schetelig MF, Bertin S, Malacrida AR, Gasperi G, Wimmer EA (2008) Fluorescent sperm marking to improve the fight against the pest insect Ceratitis capitata (Wiedemann; Diptera: Tephritidae). N Biotechnol 25:76–84
Zimowska GJ, Nirmala X, Handler AM (2009) The beta2-tubulin gene from three tephritid fruit fly species and use of its promoter for sperm marking. Insect Biochem Mol Biol 39:508–515
Acknowledgments
We extend grateful appreciation to Shelley Olson for excellent technical assistance, and to the Biotechnology Risk Assessment Program, USDA-National Institute of Food and Agriculture, for support of this project (Grant No. 2011-39211-30769).
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Schetelig, M.F., Handler, A.M. Germline transformation of the spotted wing drosophilid, Drosophila suzukii, with a piggyBac transposon vector. Genetica 141, 189–193 (2013). https://doi.org/10.1007/s10709-013-9717-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10709-013-9717-6