首页 > 核心刊物 > 遗传 2018.1 Vol 40, No.1 > 利用CRISPR/Cas9系统构建FGF21基因敲除小鼠模型

利用CRISPR/Cas9系统构建FGF21基因敲除小鼠模型

刘旭, 张平, 张晓枫, 李兴, 白宇, 贾克荣, 郭晓东, 张豪, 马晓燕, 仓明, 刘东军, 郭旭东

内蒙古大学生命科学学院,实验动物研究中心,呼和浩特 010070

 
Key Words:  FGF21, CRISPR/Cas9, Fgf21 -/-小鼠, 毛囊
 

Abstract

成纤维细胞生长因子(fibroblast growth factors, FGFs)是细胞间的多功能信号分子,调节生物体的多种生理功能。FGF21作为一种重要的调控因子,对毛囊发育及生长周期具有重要作用。为研究FGF21基因对毛囊发育及生长周期的影响及作用机制,本研究通过构建靶向敲除FGF21基因的载体,体外将Cas9 mRNA和gRNA显微注射到FVB小鼠受精卵中,在小鼠FGF21基因的第1外显子上造成移码突变,从而获得FGF21基因敲除(knock out, KO)小鼠。通过测序鉴定F0代小鼠基因型,共获得3只FGF21等位基因敲除小鼠(Fgf21 -/-);qRT-PCR和Western blotting结果表明,在Fgf21 -/-小鼠中未检测到FGF21 mRNA表达和FGF21蛋白表达;经脱毛再生及皮肤组织H&E染色发现,与野生型(wild type, WT)小鼠相比,Fgf21 -/-小鼠体重降低、器官组织未出现异常变化、毛发生长速度减慢、毛囊的直径和毛发的密度均减小。本研究利用CRISPR/Cas9技术成功构建了Fgf21 -/-小鼠模型,为后续研究FGF21基因在毛囊发育及生长周期中的功能提供了更好的动物模型。


 

Abstract

Fibroblast growth factors (FGFs) are multifunctional signal molecules between cells, regulating the various physiological functions of the organism. FGF21 is a regulatory factor of the FGF family and has been postulated to play important roles in hair follicle development and hair follicle growth cycle. To evaluate the roles of FGF21, we had established a FGF21 knockout mouse model, using the CRISPR/Cas9 technology. We had constructed a FGF21 targeting vector and microinjected it with Cas9 mRNA and gRNA into fertilized ova of FVB mice. The gRNA was designed to target the exon 1 of the endogenous mouse FGF21 gene. Three lines of Fgf21 -/- mice were obtained from these experiments, and confirmed to harbor Fgf21 -/- genotypes and null expression phenotype, using DNA sequencing, qRT-PCR and Western blotting. FGF21 mRNA and FGF21 protein were not detected in tissues of these Fgf21 -/- mice. Depilation and histochemistry analyses showed that the Fgf21 -/- mice had lower body weight, slower hair regrowth and poorer hair quantities and smaller hair follicles diameters, as compared to WT mice. The Fgf21 -/- mice reported here could provide a useful genetic model for future studies of FGF21 functions in hair follicle development and hair follicle growth cycle.


期刊简介
 
主办单位:

编辑单位:《遗传》 编辑部
主 编: 张永清
主 任: 张颖
编委阵容 :审稿专家
国内刊号:CN11-1913/R
国际刊号:ISSN 0253-9772
邮发代号: 2-810