Construction of Lep gene knockout mouse model based on CRISPR/Cas9 system
Objective We generated ob mice(C57BL/6N-Lepem1/Nifdc)with Lep gene knockout(ob/ob)using the CRISPR/Cas9 system,to establish a suitable animal model for preclinical drug evaluation for clinical diseases such as diabetes.Methods According to the principle of CRISPR/Cas9 target design,single guide RNA targeting the mouse Lep gene was designed for transcription in vitro,and microinjected with Cas9 mRNA into mouse zygotes.Mouse tail DNA was extracted and detected by polymerase chain reaction and sequencing,followed by mating of positive and wild-type mice.Blood biochemistry and liver pathology were assessed in homozygous ob mice.Results Eight positive mice were identified and a stable mouse strain was selected for further breeding.Serum triglycerides,total cholesterol,and alanine aminotransferase levels were significantly higher in homozygous ob mice than in wild-type mice,and liver pathology showed inflammatory infiltration and lipid vacuolar transformations.Conclusions We successfully established a Lep gene knockout mouse model,which will provide an important addition to the national rodent experimental animal database and an animal model for preclinical drug evaluation.
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维普
