条件性敲除软骨组织中FGF9基因小鼠模型的建立

Generation of Fgf9 conditional knockout mice in cartilage

目的:利用Cre-LoxP系统建立可调控的软骨组织条件性敲除成纤维细胞生长因子9(fibroblast growth factor,FGF9)小鼠模型。方法:构建针对小鼠Fgf9基因2号外显子的重组载体,电穿孔转染胚胎干细胞(ES细胞)。选择药物G418和Ganc,筛选阳性ES细胞并进行PCR鉴定。利用显微注射将ES细胞注入C57BL/6J小鼠囊胚,移入假孕小鼠子宫,获得含Neo的flox杂合小鼠。该小鼠去Neo后,与ColⅡ-Cre转基因小鼠杂交,并以他莫昔芬诱导,其后代软骨细胞内flox纯合、Cre阳性小鼠的flox区域被敲除。结果:Fgf9基因flox杂合子小鼠无明显异常,可稳定繁殖。该flox小鼠与ColⅡ-Cre转基因小鼠交配后,成功建立条件性敲除软骨组织中Fgf9基因小鼠模型。结论:利用Cre-LoxP系统,可成功建立Fgf9基因条件性敲除小鼠模型,为后续研究FGF9在骨软骨发育及骨关节稳态维持中的功能奠定了基础。

PURPOSE:To establish a conditional knockout mouse model of fibroblast growth factor 9(FGF9)in cartilage with Cre-LoxP system.METHODS:The recombinant vector of exon 2 of mouse Fgf9 was constructed and transfected into ES cells by electroporation.The positive ES cells were screened by G418 and Ganciclovoir and identified by PCR.ES cells were microinjected into the blastocysts of C57BL/6J mice and then transferred into the uterus of pseudopregnant mice to obtain chimeric mice.Flox region in chondrocytes of flox homozygous and Cre-positive fetuse was knocked out by hybridizing the Neo-free mice with ColⅡ-Cre mice and induced with tamoxifen.RESULTS:Fgf9 Flox heterozygous mice reproduced steadily without obvious abnormality.After mating with ColⅡ-Cre transgenic mice,the Flox mice successfully produced a conditional knockout mouse model of Fgf9 in cartilage tissue.CONCLUSIONS:Conditional knockout mice model of FGF9 was successfully established with Cre-LoxP system,which lays a foundation for further study of the function of FGF9 in osteochondral development and bone and joint homeostasis.