西南医科大学学报2024，Vol.47Issue(4)：300-305,6.DOI:10.3969/j.issn.2096-3351.2024.04.005
SNX9基因敲除小鼠模型的构建及表型分析
Construction and phenotypic analysis of SNX9 gene knockout mouse model
摘要
Abstract
Objective To construct sorting nexin 9(SNX9)gene knockout mouse model using CRISPR/Cas9 technology and analyze its phenotype.Methods sgRNA target sites were designed for exon 3～5 of SNX9 gene,and gRNA was transcribed in vitro.After mixing with mRNA encoding Cas9,F0 generation positive knockout mice were obtained by microinjection of fertilized eggs.SNX9 gene knockout homozygous(SNX9-/-)mice were obtained by breeding and genotype identification.DNA was extracted from the tail tis-sue of mice,and genotypes were determined by polymerase chain reaction(PCR)and agarose gel electrophoresis.The expressional lev-els of SNX9 and microglia marker proteins were detected by Western blot in SNX9 gene knockout mice.The reproductive rate and body weight of the SNX9 gene knockout mice were recorded.The gait of SNX9 gene knockout mice was analyzed by gait system.Hematoxylin-eosin(HE)staining was used to observe the brain structure of mouse.Results F1 heterozygous(SNX9+/-)mice mated to obtain three genotypes:wild type(SNX9+/+),heterozygous(SNX9+/-),and homozygous(SNX9-/-).DNA derived from mouse tail was used to identify mouse genotypes by PCR,and the level of SNX9 protein of SNX9 knockout mice was significantly lower than that of the wild-type mice.SNX9 gene knockout mice could grow and reproduce normally,but its reproduction rate was lower than that of wild-type mice,with normal body weight index.There were no significant differences(P>0.05)in the brain morphology and expressions of microglial marker proteins between SNX9 gene knockout mice and wild-type mice.Conclusion SNX9 gene knockout mouse model was successfully constructed using CRISPR/Cas9 techniques,which provided an experimental animal model for studying the biological func-tion and regulatory mechanism of SNX9 gene.关键词
分选连接蛋白9/CRISPR/Cas9/基因敲除/表型分析Key words
Sorting nexin 9/CRISPR/Cas9/Gene knockout/Phenotype analysis分类
医药卫生引用本文复制引用
林恒,蒋晓倩,胥元博,潘宗岳,任洋,杨朝鲜..SNX9基因敲除小鼠模型的构建及表型分析[J].西南医科大学学报,2024,47(4):300-305,6.基金项目
四川省自然科学基金(2022NSFSC0718) （2022NSFSC0718）
西南医科大学应用基础面上项目(2021ZKMS011) （2021ZKMS011）
