Objective To establish a renal carcinoma cell line A498 with CXCR4 stably silenced by using RNAi technique. Methods We designed three sequence-specific small interfering RNAs (siRNA) targeting CXCR4 gene and transfected siRNA into renal carcinoma cell line A498; the change of CXCR3 gene expression was observed by RT-PCR. The effective siRNA sequence was used to construct the recombinant plasmid, which was used to transfect A498 cells by liposome. Then the stably CXCR4 silenced cell lines were screened by using G418. RT-PCR and Westen blotting analysis were used to determine CXCR4 mRNA and protein expression. Flow cytometry was employed for determination of apoptosis in A498 cells. The invasion ability of cells was detected by transwell assay. Results Green fluorescence was seen in A498 cells transfected with recombinant shRNA plasmid. G418 screening yielded stably CXCR4-silenced A498 cell lines; RT-PCR and Western blotting analysis revealed that CXCR4 expression in CXCR4-shRNA group was significantly lower than that in the control group(P<0.05). The early apoptotic rate, late apoptotic rate, and total apoptotic rate in the CXCR4-shRNA group were significantly higher than those in the control group(P<0.05); transwell assay showed that the cell invasion ability in the CXCR4-shRNA group was significantly decreased compared with that in the control group (P<0.05). Conclusion We have successfully established an A498 cell line with CXCR4 stably silenced; the cell line has a higher apoptotic rate and lower invasion ability, which paves a way for future research.