Aim To develop novel strategies to treat familial hypercholesterolemia （FH）,we explored knocking down the endogenous hydroxymethylglutaryl CoA reductase （HMG-CoAR） expression in C57BL/6J mouse by lentivirus mediated RNAi approach in this study. Methods Three lentiviral RNAi vectors targeting sites （T1,T2 and T3） in HMG-CoA reductase gene were subject to infect H22 cell line and tail vein injection to C57BL /6J mouse after viral packaging. HMG-CoAR mRNA abundance was measured by qRT-PCR Western Blot and ELISA assay were used to detect the HMG-CoAR protein expression in infected mouse liver or plasma. Results The expression of HMG-CoAR in stably transfected H22 cells of three experimental groups was dramatically reduced in comparison to control （P<0.05）. Significantly lowing HMG-CoAR was detected in mouse liver and plasma 5 days after infection （P<0.05）. Among the three target sites,T3 was proved to be the most effective,in which the HMG-CoAR level was only 1/3 of control group. Moreover,the result of time course analysis of the inhibition effects showed that the HMG-CoAR expression was supressed at least up to 50 days in all three groups. T1 site exhibited most stable inhibition effect,yet T3 site showed more fluctuating though it was more effective than others. Conclusion Three RNAi targeted vectors were demonstrated to be able to inhibit endogenous HMG-CoAR expression in cell line and mouse. The results pave the way for lowing serum lipid in FH treatment by knocking down HMG-CoAR level.