牛c-myc基因的克隆及其在皮肤成纤维细胞中的表达  

Cloning of bovine c-myc gene and its expression in skin fibroblast cells

作　　者：肖佳佳[1] 张凤凤[1] 熊显荣[1] 张涌[1] 

机构地区：[1]西北农林科技大学动物医学院,杨凌712100

出　　处：《生物工程学报》2011年第6期963-968,共6页Chinese Journal of Biotechnology

基　　金：国家科技部转基因新品种培育重大专项~~

摘　　要：为了构建包含牛c-myc基因编码序列的重组载体,以胎牛原始生殖嵴为材料,用RT-PCR方法克隆出牛c-myc基因的编码序列,将其亚克隆至pMD19-T载体,再从酶切鉴定和测序正确的质粒上切下目的片段,定向克隆到pIRES2-AcGFP1-Nuc表达载体上,挑选序列正确的重组真核表达质粒转染牛皮肤成纤维细胞,用RT-PCR和Western blotting分别检测c-myc mRNA和蛋白的表达。结果表明,从胎牛原始生殖嵴中正确克隆了c-myc基因的全长编码序列,所构建的重组质粒能够在皮肤成纤维细胞中有效表达,为进一步研究c-myc基因功能以及下一步特定因子诱导牛体细胞转变为牛iPS细胞的研究奠定了基础。In order to construct a eukaryotic expression vector of bovine c-myc gene,the coding sequence（CDS） of c-myc gene was amplified from bovine primordial genital ridges by RT-PCR.The CDS was subcloned into pMD19-T vector,and then inserted into vector pIRES2-AcGFP1-Nuc.After confirmed by restriction enzyme digestion and sequencing,the recombined plasmid was transfected into skin fibroblast cells.RT-PCR and Western Blotting were used to detect the expression of c-myc mRNA and protein,respectively.The results show that the complete CDS of c-myc gene was cloned from fetal bovine primordial genital ridges.The eukaryotic expression vector of bovine c-myc gene was constructed and efficiently expressed in the skin fibroblast cells.The present study will lay a good foundation for further study of c-myc gene function and bovine induced pluripotent stem cells from somatic cells by defined factors.

关 键 词：牛 C-MYC基因 IPS细胞 真核表达 

分 类 号：S823[农业科学—畜牧学] Q78[农业科学—畜牧兽医]