水牛髓样分化因子88融合蛋白真核表达载体的构建及其在HEK293细胞中的表达  

Construction of Eukaryotic Expression Vector of Buffalo MyD88 Fusion Protein and its Expression in HEK293 Cells

作　　者：匡文华[1,2] 张晓茹[1,2] 成鹰[1] 杜丽[1] 雷明[1] 焦寒伟[1] 张冬琳[1] 郝永昌[1] 祁超[2] 王凤阳[1] 

机构地区：[1]海南大学农学院,海南省热带动物繁育与疫病研究重点实验室(筹),海口市动物基因工程重点实验室,海南海口570228 [2]华中师范大学生命科学学院,湖北武汉430079

出　　处：《中国畜牧兽医》2012年第4期50-54,共5页China Animal Husbandry & Veterinary Medicine

基　　金：国家转基因生物新品种培育重大专项(2009ZX08007-009B)

摘　　要：将水牛髓样分化因子88(myeloid differentiation factor 88,MyD88)cDNA全长定向克隆至pEGFP-C1真核表达载体,通过酶切和测序鉴定正确后,经脂质体介导将重组质粒转染至HEK293细胞,应用荧光显微镜、流氏细胞术和Westernblotting检测MyD88蛋白的表达。结果表明,酶切及测序结果证实重组质粒含有MyD88全长CDS序列,融合蛋白读码正确;荧光显微镜、流式细胞术和Western blotting检测证实,水牛EGFP-MyD88融合蛋白真核表达载体能够在HEK293细胞表达。本研究成功构建了pEGFP-C1-MyD88真核表达载体,并使其在HEK293细胞中获得表达,为进一步研究该基因的生物学功能奠定基础。The recombinant vector constructed by gene recombinant technology was analyzed by restriction enzyme digestion and DNA sequencing.The recombinant vector was transfected into HEK293 cells with liposome for transient expression.The expression of fusion protein buffalo MyD88 was identified by fluorescent microscope,flow cytometry and Western blotting.The recombinant plasmid proved successful by restriction enzyme digestion and DNA sequencing.The expression of fusion protein was shown by fluorescent microscope,flow cytometry and Western blotting.It is concluded that the recombinant eukaryotic vector pEGFP-C1-MyD88 was successfully constructed and the fusion protein successfully expressed in HEK293 cells.

关 键 词：水牛 髓样分化因子88 EGFP 融合蛋白 真核表达 

分 类 号：S786[农业科学—林学]