心肌锚定重复序列蛋白基因心脏特异敲除小鼠模型构建与鉴定

基础医学与临床 ›› 2011, Vol. 31 ›› Issue (11): 1217-1222.

心肌锚定重复序列蛋白基因心脏特异敲除小鼠模型构建与鉴定

许家林¹,李艳凤²,朱大海³

1. 中国医学科学院基础医学研究所
2. 哈尔滨工业大学分子与细胞发育生物学实验室
3. 中国医学科学院北京协和医学院基础医学研究所

收稿日期:2011-09-16 修回日期:2011-09-20 出版日期:2011-11-05 发布日期:2011-11-02
通讯作者: 朱大海 E-mail:dhzhu@pumc.edu.cn

Constriction and identification of mice model with cardiac specific disruption of cardiac ankyrin repeat protein

Received:2011-09-16 Revised:2011-09-20 Online:2011-11-05 Published:2011-11-02

摘要/Abstract

摘要： 摘要: 目的构建心肌锚定重复序列蛋白（CARP）基因心脏特异敲除小鼠模型，为研究CARP基因与心脏疾病奠定基础。方法构建CARP基因条件打靶载体PL253-CARP-LoxP电转入小鼠胚胎干细胞（ES）细胞，G418和GANC药物筛选得到阳性细胞克隆，Southern blot确定CARP条件打靶载体转染成功。胚胎注射阳性克隆ES细胞入小鼠囊胚，获得嵌合体小鼠与心脏特异启动子α-MHC调节Cre小鼠（α-MHC-Cre）杂交，得到CARP基因心脏特异敲除小鼠（CARP-KO）。结果基因型鉴定获得6只嵌合体小鼠，与α-MHC-Cre小鼠杂交获得CARP-KO小鼠，半定量PCR和Western blot确定CARP基因在CARP-KO小鼠心脏缺失。该小鼠早期胚胎发育和个体生长、心脏发育正常，但心脏肥厚标志基因β-MHC和ANF显著升高。结论：成功构建CARP基因心脏特异敲除小鼠，该基因缺失对小鼠胚胎早期发育、生长和心脏发育没有显著影响。

关键词: 心肌锚定重复序列蛋白, 基因打靶, 心肌肥厚

Abstract: Abstract: Objective To establish the mice model with cardiac disruption of cardiac ankyrin repeat protein (CARP) and to provide reliable tools for studying the function of CARP gene in heart related diseases. Methods Construct the conditional target plasmid of PL253-CARP-LoxP and transfect to 129 embryonic stem cells. Positive clones selected by antibiotics of G418 and GANC were determined by Southern blot to confirm conditional disruption of CARP gene, and then were injected to the blastula. Chimeras were got and crossed with α-MHC-Cre transgenic mice, which expressed recombinase specifically in heart, and then the mice with cardiac deficiency of CARP gene (CARP-KO) will get. Results Six chimeras were got, and CARP-KO mice were got after crossing with α-MHC-Cre mice. CARP gene was disrupted in the heart of CARP-KO mice by semi-quantitative PCR and western blot. These mice displayed normal in embryonic development, body growth, and heart development. However the expression of hypertrophic markers of β-MHC and ANF increased significantly in CARP-KO mice. Conclusion: The mice model with specific disruption of CARP gene was generated successfully. The cardiac disruption of CARP gene did not interfere the development and growth of mice significantly, either for the heart development.

Key words: cardiac ankyrin repeat protein, gene targeting, hypertrophy

许家林李艳凤朱大海. 心肌锚定重复序列蛋白基因心脏特异敲除小鼠模型构建与鉴定[J]. 基础医学与临床, 2011, 31(11): 1217-1222.

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