Objective To analyze the expression of miR-125b-5p in lung adenocarcinoma (LUAD), and predict its target genes and prognostic value by bioinformatics analysis. Methods Tissue and cell chips of cisplatin resistance-related LUAD in Gene Expression Omnibus (GEO) database were obtained, and differentially expressed miRNAs were screened by R software. The miRBase database was used to analyze conservativeness, and the dbDEMC3.0 database was used to evaluate the expression level. The target genes were predicted by TargetScan, miRDB and miRTarBase databases, and were analyzed for functional term enrichment with the DAVID database. Combined with the survival information of LUAD patients in the Cancer Genome Atlas (TCGA) database, prognostic analysis was performed. Quantitative Real-time PCR (qRT-PCR) detection technology was used to detect the expression level of miR-125b-5p in human normal bronchial epithelial cell line 16HBE, LUAD cell line A549, and cisplatin-resistance cell line A549/DDP. Results miR-125b-5p, closely related to cisplatin resistant of LUAD, was obtained by analysis of the chip data. The sequence of miR-125b-5p was highly conserved among species, and miR-125b-5p was significantly down-regulated in various tumors including LUAD. Fifty-four target genes of miR-125b-5p were predicted. The results of enrichment analysis showed that the target genes of miR-125b-5p were mainly involved in the regulation of gene expression, cellular macromolecule biosynthetic process, and mainly enriched on MicroRNAs in cancer, Protein processing in endoplasmic reticulum, and HIF-1 signaling pathway. Survival analysis indicated that miR-125b-5p was significantly associated with poor prognosis of patients with LUAD. qRT-PCR results showed that the expression level of miR-125b-5p in A549 was significantly lower than in 16HBE (P=0.008). Compared to parental cell, the expression level of miR-125b-5p was significantly down-regulated in A549/DDP cell (P=0.023). Conclusion miR-125b-5p was abnormally expressed in LUAD, and its target genes involved in multiple biological processes and signal pathways, which might be used as a new biomarker for prognosis in LUAD.