利用CRISPR/Cas9构建R778L类型肝豆状核变性疾病小鼠模型

基础医学与临床 ›› 2018, Vol. 38 ›› Issue (12): 1702-1707.

利用CRISPR/Cas9构建R778L类型肝豆状核变性疾病小鼠模型

朱基彦¹,瞿思遥¹,张祝琴²,刘德培²

1. 中国医学科学院基础医学研究所
2. 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:2018-09-17 修回日期:2018-10-24 出版日期:2018-12-05 发布日期:2018-11-23
通讯作者: 刘德培 E-mail:liudp@pumc.edu.cn
基金资助:
国家重点研发计划;中国医学科学院医学与健康科技创新工程

Construct R778L type hepatolenticular degeneration mouse model with CRISPR/Cas9 system

Received:2018-09-17 Revised:2018-10-24 Online:2018-12-05 Published:2018-11-23

摘要/Abstract

摘要： 目的利用CRISPR/Cas9介导的基因编辑系统构建针对人肝豆状核变性R778L突变类型的R780L突变小鼠。方法通过BLAST比对人和小鼠中ATP7B基因和蛋白的序列，证明二者保守性。通过CRISPR/Cas9系统进行小鼠受精卵显微注射，并对小鼠肝豆状核变性症状进行病理和生理学检测。结果人和小鼠中ATP7B基因和蛋白的序列高度保守。通过CRISPR/Cas9技术成功获得纯合的R780L突变小鼠，该小鼠存在明显的肝脏铜离子淤积以及血清ALT、AST的升高且未发现有可检测出的脱靶效应。结论成功利用CRISPR/Cas9介导的基因敲入系统构建了针对人肝豆状核变性R778L类型的R780L突变小鼠模型。

关键词: 肝豆状核变性, CRISPR/Cas9, 点突变R778L, 小鼠模型

Abstract: Objective R780L mutant mice directed against the human hepatolenticular degeneration R778L mutation type were constructed using a CRISPR/Cas9-mediated gene editing system. Methods The conserved nature of the ATP7B gene and protein in humans and mice was confirmed by BLAST. Microinjection of mouse fertilized eggs was carried out by the CRISPR/Cas9 system, and pathological and physiological tests were performed on the symptoms of hepatolenticular degeneration mouse model. Results The sequences of ATP7B gene and protein between humans and mouse were highly conserved. The homozygous R780L mutant mouse were successfully obtained by CRISPR/Cas9 system, which showed significant liver copper ion deposition and elevated serum ALT and AST and no detectable off-target effects were observed. Conclusion Successfully constructed a R780L mutant mouse model for human hepatolenticular degeneration R778L type using CRISPR/Cas9-mediated gene knock-in system.

Key words: Wilson's disease, CRISPR/Cas9, Point mutation R778L, Mouse model

朱基彦瞿思遥张祝琴刘德培. 利用CRISPR/Cas9构建R778L类型肝豆状核变性疾病小鼠模型[J]. 基础医学与临床, 2018, 38(12): 1702-1707.