Characterization of the Drosophila suzukii β2-tubulin gene and the utilization of its promoter to monitor sex separation and insemination

The Drosophila melanogaster beta 2-tubulin gene (Dm beta 2t) controls the function of microtubules in the testis and sperm, and has been evaluated for use in biocontrol strategies based on the sterile insect technique, including sexing and the induction of male sterility. The spotted-wing Drosophila (Drosophila suzukii) is native to eastern Asia but has spread globally as an invasive pest of fruit crops, so biocontrol strategies are urgently required for this species. We therefore isolated the beta 2tubulin ortholog Ds-beta 2t from the USA laboratory strain of D. suzukii and confirmed the presence of functional motifs by aligning orthologs from multiple insects. The developmental expression profile of Ds-beta 2t was determined by RT-PCR using gene-specific primers and was similar to that of Dm-beta 2t. We then isolated the Ds beta 2t promoter and used it to generate transgenic strains expressing a testis-specific fluorescent protein starting from the thirdinstar larvae. Efficient sexing was achieved based on fluorescence detection, and the transgenic males showed a similar survival rate to wild-type males. Fluorescence imaging and PCR were also used to confirm the insemination of wild-type females by transgenic males. We therefore confirm that D. suzukii strains expressing fluorescent markers under the control of the Ds-beta 2t promoter can be used for sexing and the confirmation of mating, and we discuss the wider potential of the Ds-beta 2t promoter in the context of genetic control strategies for D. suzukii.