Routine chemotherapy as an important treatment mode often can not be effective because of multidrug resistance (MDR). MicroRNA (miRNA) modulates the expression of a great number of genes, including MDR. In this study, the expression of miR-1284 was reduced in gastric cancer (GC) tissue specimens with metastasis and in vincristine-resistant (VCR) GC SGC7901 cells (SGC-7901/VCR) compared to that in the controls. Recombinant lentiviral vectors with miR-1284 led to the overexpression of miR-1284 mRNA and reversed the chemoresistance of SGC7901/VCR cells, promoted cell cycle arrested at the G0/G1 phase, accelerated drug-induced apoptosis, and decreased migration and invasiveness of SGC-7901/VCR. In addition, the overexpression of miR-1284 sensitized tumors to chemotherapy in vivo. Our data provide combined evidence that miR-1284 can heighten the expression of MYC and reduce the expression of JUN, MMP12, and EIF4A1 that was the direct target. In conclusion, miR-1284 can function as a new regulator to reduce GC MDR cells by targeting EIF4A1.