首页|采用Cre-loxP技术构建肝脏特异性Trappc11基因敲除小鼠模型

采用Cre-loxP技术构建肝脏特异性Trappc11基因敲除小鼠模型

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目的 采用Cre-loxP系统构建肝脏组织特异性Trappc11基因敲除小鼠(Trappc11flox/flox-Alb-Cre+/-)模型以探究脂肪肝的发病机制.方法 用带有loxP位点的Trappc11纯合子小鼠(Trappc11flox/flox)和带有Alb-Cre的转基因小鼠(Alb-Cre+/-)进行杂交,对其子代进行基因型鉴定.筛选出Trappc11fllox/+-Alb-Cre+/-小鼠,将其与Trappc11fox/flox小鼠进一步杂交可获得Trappc11肝脏特异性敲除鼠.将3~4周龄的Trappc11flox/flox-Alb-Cre+-、Trappc11flox/+-Alb-Cre+/-和Trappc11flox/flox-Alb-Cre-/-小鼠各3只,提取各组织脏器中的DNA,检测Trappc11的敲除效率和特异性.然后,在mRNA和蛋白质水平进一步验证.于第4周开始监测摄食量和体重等指标.结果 我们成功繁育出肝脏特异性Trappc11基因敲除鼠,敲除心、脾、肺、肾、胰腺、肌肉、脂肪和脑组织中该基因表达未受影响,mRNA水平敲除效率达85%以上且TRAPPC11蛋白水平下降.该敲除鼠目前生长状态良好,饮食饮水正常,可用于后期繁殖与机制研究.结论 已经构建成功的Trappe11flox/flox-Alb-Cre+/-鼠将为探索Trappc11在肝脏中的生理病理作用提供在体模型.
Establishment of a liver-specific Trappc11 gene knockout mouse model using the Cre-loxP system
Objective A liver-specific Trappc11 gene knockout mouse model(Trappc11flox/flox-Alb-Cre+-)was established using the Cre-loxP system to investigate the pathogenesis of fatty liver disease.Methods Trappc11 homozygous mice with a loxP site(Trappc11flox/flox)were crossed with Alb-Cre transgenic mice(Alb-Cre+/-),and the genotypes of the offspring were identified.Progeny with the Trappc11/flox-Alb-Cre+-genotype were selected,and liver-specific knockout mice were obtained through further breeding with Trappc11flox/flox mice.Three groups of mice,including Trappc11flox/flox-Alb-Cre+/-、Trappc11flox/+-Alb-Cre+/-and Trappc11flox/flox-Alb-Cre-,aged 3-4 weeks,were selected,DNA was extracted from tissue samples to assesst the knockout efficiency and specificity of Trappc11.with further validation at the mRNA and protein levels.By the forth week,parameters such as food intake and body weight were measured.Results We successfully generated liver-specific Trappc11 gene knockout mice.Although the expression of Trappc11 gene was not affected in various tissues,including the heart,spleen,lung,kidney,pancreas,muscle,fat and brain,our model achieved a knockout efficiency of over 85%at the mRNA level,significantly decreasing the TRAPPC11 protein level.The knockout mice are currently in good health,exhibiting normal growth,diet and water intake,making them suitable for subsequent breeding and mechanistic studies.Conclusion The successfully constructed Trappc11flox/flox-Alb-Cre+/-mice provide a valuable in vivo model for investigating the physiological and pathological roles of Trappc11 in the liver.

Trafficking Protein Particle Complex Subunit 11Cre-loxP recombinase systemTissue specific knockout

郑帆帆、李霞、高枫、李梦瑶、杨彦玲、李优磊

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716000 陕西 延安大学医学院

延安市肥胖防治研究重点实验室,延安大学附属医院内分泌代谢科

转运蛋白颗粒复合物亚基11 Cre-loxP重组酶系统 组织特异性敲除

陕西省教育厅科学研究项目陕西省自然科学基础研究计划延安大学博士科研启动项目

22JK06132024JC-YBQN-0949YDBK2021-07

2024

肝脏
上海市医学会

肝脏

CSTPCD
影响因子:0.71
ISSN:1008-1704
年,卷(期):2024.29(9)