牛myf6基因克隆及在成纤维细胞中的表达

Cloning of bovine myf6 gene and expression of gene in fibroblasts

克隆了牛的myf6基因并构建真核表达载体pIRES2-EGFP-myf6,用脂质体技术转染鲁西黄牛成纤维细胞,通过G418筛选出稳定转染的细胞株,利用Western印记、Real-time PCR技术检测myf6对成纤维细胞的影响。结果表明,与对照组相比,稳定转染后的成纤维细胞myf6蛋白和mRNA的表达量提高(P<0.01),肌肉肌酸激酶基因mRNA表达量升高(P<0.05),肌球蛋白轻链基因的mRNA表达量也提高(P<0.01)。细胞形态观察显示转染后的成纤维细胞未融合为肌管。由此可知,牛myf6基因可以在成纤维细胞中表达并且有促进成纤维细胞向肌肉细胞分化的功能。

myf6 gene was a member of the MRFs family.It controlled the cell proliferation and differentiation.In this article,genes of the myf6 were cloned and eukaryotic expression vector pIRES2-EGFP-myf6 was constructed.Liposome techinigue was used to transfect Luxi cattle fibroblasts,and by G418 selection,a stable transfected cell line was gained.The effects of fibroblasts were analyzed by Western blot and Real-time PCR.Compared with the control group,the results showed that after the fibroblasts transfected by plasmid,protein and mRNA expression levels of myf6 both increased（P0.01）,mRNA expression of the muscle creatine kinase genes increased（P0.05）,and mRNA expression of myosin light chain gene had increased（P0.01）.Morphology observations indicated that fibroblasts did not fuse to myotubes.The results suggested that myf6 gene was expressed in fibroblasts and promoted fibroblasts differentiation.