Objective To explore the aberrantly expressed genes in hepatocellular carcinoma (HCC) and their relationship with prognosis of HCC through bioinformatics analysis. Methods Five datasets related to HCC were selected from the GeneExpression Omnibus database to explore differentially expressed genes (DEGs), followed by further gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The co-upregulated genes CNIH4 and TOMM40 were selected to explore the differences in their expressions in HCC tissues and normal tissues, and to explore the relationship between their expressions and the 5-year survival of patients by using TCGA database. Tissues and paraneoplastic tissues of eight cases of HCC who underwent surgery at the Guangdong Second Provincial General Hospital were collected to verify the expression differences of CNIH4 and TOMM40L mRNA. Results A total of 25 up-regulated genes and 21 down-regulated genes were identified in this study. The results of GO analysis and KEGG analysis indicated that DEGs were mainly related to catabolism, cell division, DNA replication and repair. The results of TCGA database analysis showed that the expression of up-regulated genes CNIH4 mRNA and TOMM40L mRNA were up-regulated in HCC tissues as compared with normal tissues (P<0.05) and that the 5-year survival of patients in the high expression group was worse than that in the low expression group (P<0.05). The results of clinical samples showed that CNIH4 mRNA and TOMM40L mRNA were up-regulated in HCC tissues as compared with paraneoplastic tissues. Conclusion CNIH4 and TOMM40L genes are up-regulated in HCC tissues, and their high expressions are associated with poor prognosis, and may be potential biomarkers and prognostic indicators for HCC.