Ipl 41 medium

BmN-4 cells (provided by Chisa Yasunaga-Aoki, Kyushu University, and maintained in our laboratory)⁶ were cultured at 27°C in IPL-41 medium (Applichem) supplemented with 10% fetal bovine serum. VF and VF-MLV cells (established and maintained in our laboratory) were cultured at 27°C in IPL-41 medium (Applichem) supplemented with gamma ray-treated 10% fetal bovine serum.⁵ (link)

BmN4 cells were cultured at 27°C in IPL-41 medium (AppliChem) supplemented with 10% fetal bovine serum. For immunoprecipitation experiments, a total of 5–7.5 µg of plasmid and dsRNAs were transfected into BmN4 cells (2–2.5 × 10⁶ cells per 10 cm dish) with X-tremeGENE HP DNA Transfection Reagent (Sigma), and the transfected cells were harvested after 4 d. For cotransfection experiments, the second transfection was performed 2 d after the first transfection, and the cells were harvested after an additional 4 d. For immunofluorescence experiments, a total of 0.4 µg of plasmid and dsRNAs were transfected into BmN4 cells (4–6 × 10⁴ cells per glass bottom 35 mm dish) with X-tremeGENE HP DNA Transfection Reagent (Sigma), and cells were fixed 4 d later. For Supplemental Figure 2, transfection was repeated 2 d later and cells were fixed 4 d after the second transfection. For library preparation, 5 µg of dsRNAs were transfected into BmN4 cells (8 × 10⁵ cells per 10 cm dish) with X-tremeGENE HP DNA Transfection Reagent (Sigma) every 3 d four times. dsRNA preparation was described previously (Izumi et al. 2020 (link)). Template DNAs were prepared by PCR using primers containing T7 promoter listed in Supplemental Table 1.