Recombinant Baculovirus Expression of OfCad in Sf9 Cells

Spodoptera frugiperda Sf9 cell lines [36 (link)] were cultured in Sf-900™ III SFM medium (Thermo Fisher Scientific, Waltham, MA, USA) at 27 °C with a density of 5 × 10⁶ cells/mL and subcultured every 3 d. Log phase cells were used for infection and were seeded at a density of 2 × 106 cells mL−1, and transfected with P3 recombinant baculovirus stocks. By following the operation manual of Bac-to-Bac baculovirus expression system (Thermo Fisher Scientific, Waltham, MA, USA), purified pFastBac™ HTA-OfCad plasmids were transformed into E. coli DH10Bac cells to generate recombinant bacmids. Purified recombinant bacmids were used to infect Sf9 by using the FuGENE HD transfection reagent (Promega, Madison, WI, USA). P3 viral stocks were stored at 4 °C for further use. Negative control used in the current study includes cells transfected with empty bacmid (EB) and the non-transfected cells (Sf9).