Construction of SLC19A2 knockout mice based on CRISPR/Cas9 techno-logy and its phenotypic validation
Objective To construct a purebred mouse model of SLC19A2 knockout using CRISPR/Cas9 technology and validate its phenotype.Methods The gRNA vectors were designed,and the constructed gRNA vectors and Cas9 vectors were co-injected into fertilised eggs after in vitro transcription,and six(three females and three males)positive F0 generation mice were obtained after embryo transfer.Twelve(six males and six females)SLC19A2 knockout purebred mice[SLC19A2-/-mice(C57BL/6)]were obtained by breeding.Twelve wild-type C57BL/6 mice and twelve SLC19A2-/-mice(six males and six females)each at eight weeks of age were set up as the wild-type group and the knockout pureblood group,respectively.Mouse genotypes were identified by PCR amplification gel electrophoresis;SLC19A2 mRNA expression in pancreatic and liver tissues was detected by real-time fluorescence quantitative PCR,and THTR-1 protein expression in pancreatic and liver tissues was detected by Western blot.Results SLC 19A2 mice were successfully identified by PCR amplification gel electrophoresis.The expression levels of SLC 19A2 mRNA and THTR-1 protein in pancreatic tissues and liver tissues of the knockout pureblood group were lower than those of the wild-type group,and the differences were statistically significant(P＜0.05).Conclusion SLC19A2 knockout mice were successfully constructed based on CRISPR/Cas9 technology.
万方数据
维普
