Objective To knock out the Abcb1 gene of rat,and establish the Abcb1 humanized rat model based on the Abcb1 knock out rat. Methods The animal model was established using BAC and CRISPR/Cas9 technology,and was analyzed by PCR, RT-PCR and real-time PCR. Results Establishing a rat model expressing human Abcb1 stably by transfer the 153 kb BAC containing human Abcb1 promoter and cDNA into rat genome, and establishing the Abcb1 knock out rat at the same time. Establishing the Abcb1 humanized model by crossing these two strains together. The expression pattern of Abcb1 in Abcb1 humanized rat is different from the wild type rat. The Abcb1 humanized model express not only the human Abcb1 gene but also has similar expression pattern as human. Conclusions The Abcb1 knock out rat and the Abcb1 humanized rat were successfully established, and this model is close to human concerning about the drug metabolism related to Abcb1.