We found that the transcription factor Krüppel-like factor 8 (KLF8) was highly expressed in gastric cancer tissues and cell lines compared with adjacent noncancerous regions and gastric epithelial mucosa cells. We employed a lentivirus-mediated RNAi technique to knockdown KLF8 expression in gastric cancer cell line SGC7901 and observed its effects on cell growth in vitro and in vivo. Knockdown of KLF8 inhibited SGC7901 cell proliferation, promoted cell apoptosis, inhibited the tumorigenicity of SGC7901 cells, and significantly decreased tumor growth when the cells were injected into nude mice. These results indicated that overexpression of KLF8 may influence the biological behavior of SGC7901 gastric cancer cells. Knockdown of KLF8 expression by lentivirus-delivered siRNA may be useful as a therapeutic agent for the treatment of gastric cancer.