RNA transfection

Lonza Cell Line Nucleofector kit V was used to introduce endoribonuclease-prepared siRNA targeting ORP5 (Cat no. EHU085181-20UG) and ORP8 (Cat no. EHU070271-20UG) in HepG2 cells. Alternatively, Lipofectamine RNAiMAX (Life Technologies) was used to introduce siRNAs to the cells following the manufacturer’s protocol. The esiRNA targeting Renilla Luciferase (RLuc; Cat no. EHURLUC-20UG) was used as the control. Each reaction contains 1 × 10⁶ cells and 30 nM siRNA. The H-022 program for high viability was selected. After the program, HepG2 cells were seeded at a density of 200,000 cells per 35-mm dish for 72 h before imagining.