Use of CRISPR/Cas9 system for establishment and characterization of Apoe gene knockout mice model
Objective The CRISPR/Cas9 system was utilized to generate an Apoe knockout mice model to support further investigations of the role of Apoe in lipid metabolism and atherosclerosis.Methods Two single guide RNAs designed for Apoe in C57BL/6J mice were co-injected with Cas9 mRNA into fertilized eggs,followed by transplantation into ICR recipient mice to obtain F0 generation mice.KO mice were identified by polymerase chain reaction(PCR)screening of tail DNA.Apoe mRNA expression in various tissues was assessed by quantitative real-time PCR and lipid indexes were measured in serum samples.Lipid accumulation in the inner lining of aortic vessels was detected by oil red O staining.Results PCR and sequencing confirmed the successful construction of Apoe KO mice(C57BL/6-Apoeem1/Nifdc).Apoe mRNA levels were significantly reduced in the liver,brain,spleen,kidney,and lung tissues of Apoe KO homozygous mice(Apoe-/-),as shown by reverse transcription quantitative real-time PCR.Serum total cholesterol and low-density lipoprotein cholesterol levels were increased in Apoe-/-mice,and high-density lipoprotein cholesterol levels were decreased in male Apoe-/-mice.Extensive lipid plaques were observed in the inner lining of the arteries in Apoe-/-mice compared with WT mice,under normal chow consumption conditions.Conclusions This study successfully established an Apoe KO mice model exhibiting a typical abnormal lipid metabolism phenotype with arterial lipid accumulation,even without a high-fat diet intervention.This work provides background data for the Apoe KO mice resource and a new model for the study of abnormal lipid metabolism.
万方数据
维普
