Objective To develop apolipoprotein E ( ApoE) gene-knockout rabbits, investigate their phenotypic characteristics and reproductive performance, and establish a breeding technology system for ApoE geneknockout rabbits. Methods ApoE gene-knockout rabbits were constructed using CRISPR/ Cas9 gene-editing technology with embryo transplantation. Healthy ApoE gene-knockout rabbits were screened by genotyping to serve as breeding stock and their selection and expansion were conducted by inbreeding or backcrossing. Phenotypic analysis included routine blood tests, lipid biochemical assays, and pathological examination. Reproductive performance was also evaluated to select genetically stable ApoE gene-knockout rabbits with distinct phenotypes. Results An ApoE ^-/-rabbit population was successfully established. Under a normal diet, total cholesterol and low-density lipoprotein cholesterol levels were significantly higher in ApoE ^-/- than in New Zealand rabbits ( WT) ( P< 0. 01), exhibiting pathological features of lipid metabolism disorder and atherosclerosis. There were no significant differences in reproductive performance and growth development between ApoE ^-/- and New Zealand rabbits (WT). Conclusions This study successfully produced and selected genetically stable and phenotypically distinct ApoE gene-knockout rabbits, establishing an effective breeding technology system. These rabbits provide an important animal model for studying the mechanisms of cardiovascular and related metabolic diseases.