Establishment of transgenic Drosophila suzukii lines that express phiC31 integrase and carry the sepia gene as a marker for transformation.

Many eye colour mutants have been identified in Drosophila melanogaster. Mutations in the sepia gene result in brown eyes due to a lack of PDA synthase, which is essential for production of the red drosopterin eye pigment. We previously used CRISPR/Cas9 to target the PDA synthase gene to establish sepia mutant strains for Drosophila suzukii (Matsumura) (Diptera: Drosophilidae), an invasive global pest of soft skinned fruits. The fecundity and fertility of some of the sepia mutant strains were similar to wild-type. The goal of this study was to determine if the sepia gene could be used as a marker to identify transgenic D. suzukii. By using the sepia gene as a marker, we successfully developed lines expressing Streptomyces phage phiC31 integrase in the germline. For most of these lines, hemizygotes exhibited complete rescue of the sepia eye colour and relatively high levels of phiC31 RNA in ovaries. In contrast, lines with partial rescue showed low levels of sepia RNA in heads and phiC31 RNA in ovaries. These findings suggest that the sepia gene is an effective marker for D. suzukii transgenesis and its relatively small size (1.8 kb) makes it advantageous when assembling large gene constructs. The phiC31 integrase lines established in this study should serve as a valuable resource for future genetic research in D. suzukii, including the further development of strains for genetic biocontrol.