In the present study, we prepared the small interfering RNA (siRNA) complexes with polyamidoamine (PAMAM) dendrimer (G3) conjugate with α-cyclodextrin (α-CDE (G3)), and examined the physicochemical properties, serum resistance, in vitro RNAi effects on endogenous gene expression, cytotoxicity, interferon response, hemolytic activity, cellular association and intracellular distribution. In addition, these results were compared to the siRNA complexes with the commercial transfection reagents such as linear polyethyleneimine (PEI), Lipofectamine™2000 (L2) and RNAiFect™ (RF). α-CDE (G3) interacted with siRNA, and suppressed siRNA degradation by serum. The siRNA complex with α-CDE (G3) showed the potent RNAi effects against Lamin A/C and Fas expression with negligible cytotoxicity and hemolytic activity, compared to those of the transfection reagents in Colon-26-luc cells and NIH3T3-luc cells. Cell-death patterns induced by siRNA polyplexes with α-CDE (G3) and PEI were different from siRNA lipoplexes with L2 and RF. α-CDE (G3) delivered fluorescent-labeled siRNA to cytoplasm, not nucleus, after transfection in NIH3T3-luc cells. Taken together, α-CDE (G3) could be potentially used as a siRNA carrier to provide the RNAi effect on endogenous gene expression with negligible cytotoxicity.