Furthermore, through the negative selection screen, we identified several novel knockout combinations for kinases and metabolism‐associated genes implicated in HCC tumourigenesis, such as Raf1_Pkm2 and Raf1_Top2α, Raf1_Alox5, Raf1_Drd1, Raf1_Nqo1 and Raf1_Rxra, which belong to the Raf1‐associated gene pairs, and showed that hepa1‐6 cells are sensitive to Raf1 perturbation. Here, RAF1 is linked to hepatocellular carcinoma.