Objective: This study aimed to investigate the roles of MRPL27 in survival from cholangiocarcinoma patients in The Cancer Genome Atlas (TCGA) database. Methods: In TCGA-CHOL profile, MRPL27 gene expression and clinical data were obtained. Cox regression models were used to evaluate the potential links between MRPL27 and cholangiocarcinoma survival. Enrichment analysis of MRPL27 was conducted in Metascape and Gene Set Enrichment Analysis (GSEA) databases. Results: 36 cholangiocarcinoma patients were included in this analysis. MRPL27 mRNA was significantly upregulated in tumor tissues in cholangiocarcinoma patients including intrahepatic, distal and hilar/perihilar cholangiocarcinoma cases (all p < 0.01). Cholangiocarcinoma patients with high MRPL27 had worse overall survival (OS) and disease-free survival (DFS) compared to those with low MRPL27 (all p < 0.05). Univariate and multivariate Cox models indicated that MRPL27 should be a risk factor for the OS and DFS in cholangiocarcinoma patients (both p < 0.01). Bioinformatic analysis revealed that MRPL27 mainly involved in the processes of mitochondrial translation elongation, respiratory electron transport, ATP synthesis, and inner mitochondrial membrane organization. No mutations of MRPL27 were screened in cholangiocarcinoma patients. Conclusion: Upregulated in tumors, MRPL27 contributes to unfavorable survival in cholangiocarcinoma patients.