Gastric cancer (GC) is one of the most common malignant tumors with a high mortality rate. Reversing the multi-drug resistance (MDR) of GC offers the potential for significant enhancement of the effect of chemotherapy and improvement of prognosis. Aberrant microRNA expression can attribute to the pathogenesis of GC. However, the effects of microRNA (miR)-195-5p on the MDR of GC cells remains to be fully elucidated. In the present study, the effect of miR-195-5p in regulating the MDR of GC cells was investigated. Reverse transcription quantitative-polymerase chain reaction was used to analyze the levels of miR-195-5p in GC cells. Western blot analysis was performed to analyze the protein levels of ZNF139, P-gp, BCL-2 and MRP1. The chemosensitivity of GC cells was determined by MTT. The results showed that the expression of miR-195-5p was decreased in poorly differentiated GC tissues with a higher chemosensitivity. The overexpression of miR-195-5p promoted the chemosensitivity of GC cells. Bioinformatics analysis indicated that Zing finger 139 (ZNF139) was a target of miR-195-5p. miR-195-5p negatively regulated the expression of ZNF139 by binding to its 3-untranslated region. The silencing of ZNF139 promoted the chemosensitivity of GC cells, and the downregulation of ZNF139 reversed the effect of miR-195-5p inhibitor on the chemosensitivity of GC cells. In conclusion, miR-195-5p regulated the MDR of GC cells via targeting ZNF139.