The aim of the present study was to construct the eukaryotic expression vector pcDNA3.1/15-PGDH. The vector was used to transfect mouse murine forestomach carcinoma (MFC) cancer cells and observe the effects of 15-hydroxyprostaglandin dehydrogenase (15-PGDH) on the proliferation of MFC. pcDNA3.1/15-PGDH was constructed using gene recombination technology and the vector was used to transfect MFC cells to build a stable transfected cell strain. The expression levels of 15-PGDH in the transfected cells were detected using reverse transcription polymerase chain reaction. Optical Density (OD) values were determined using an MTT assay and used to draw cell growth curves. The effects of 15-PGDH on the proliferation of MFC were observed using a clone formation experiment. Following successful transfection by 15-PGDH, the relative expression levels of 15-PGDH in the MFC/15-PGDH cells were significantly higher (1.06 +/- 0.08) (P<0.01) compared with the empty plasmid-transfected group (0.22 +/- 0.01) and the untransfected group (0.21 +/- 0.01). Following transfection by 15-PGDH, cell growth was markedly inhibited. The MTT results showed that on days 4, 6 and 8, the 15-PGDH-transfected group had a low OD on average, which was significantly different (P<0.05) from the empty plasmid-transfected group or the untransfected group. The 15-PGDH-transfected group had a plating efficiency of 18%, and compared with the untransfected group (63%) and the empty plasmid-transfected group (59%), clone formation was significantly inhibited (P<0.01). Results of the present study indicate that transfection by 15-PGDH may significantly inhibit the proliferation and clone formation of MFC cells.