Yeast Two-Hybrid Screening of Viral Protein Interactions

To examine the interaction among RdFV CP, HongrES1, and RGDV P8, the DUALmembrane starter kit (Dualsystems Biotech, P01201-P01229) was used according to the manufacturer’s instructions. The full-length ORFs of RdFV CP and RGDV P8 were separately inserted into bait vector pDHB1 to generate pDHB1-CP and pDHB1-P8, and the full-length ORFs of HongrES1, RdFV CP, RSMV G, and RDV P8 were separately inserted into prey vector pPR3-N to generate pPR3-N-HongrES1, pPR3-N-CP, pPR3-N-G and pPR3-N-P8 (RDV). The bait and prey were co-transformed the yeast strain NMY51. The pLargeT/p53 interaction served as a positive control, and the pDHB1/pRR3N served as a negative control. Transformants were subsequently screened on the QDO (SD/-Trp-Leu-His-Ade/X-Gal) culture medium, and β-galactosidase activity was detected in Z buffer with X-Gal. The primers used in Y2H were shown in Supplementary Table 1.