摘要
为在原核细胞中表达TGF β1单体蛋白 ,用基因重组技术构建pBV2 2 0 TGF β和 pQE TGF β两种原核表达载体 ,分别在M15和DH5α大肠杆菌中进行表达 ,然后利用SephacrylS 10 0凝胶层析及Ni+ NTA琼脂柱纯化TGF β1单体。经酶切鉴定及测序结果证明pBV2 2 0和pQE30载体上成功地插入了TGF β1成熟肽基因片段。pBV2 2 0 TGF β和pQE TGF β两种原核表达载体在大肠杆菌中均得到了高效表达 ,表达量约占全菌蛋白的 15 %和 2 0 % ,表达的TGF β1单体蛋白经纯化后进行SDS PAGE电泳 ,均显示了一条蛋白带 ,分子量分别为 13kD和 15kD左右。
To further understand the function and biological activity of TGF β 1, and to provide the basis for the production of biologically active TGF β 1 protein, TGF β 1 cDNA was cloned into procaryotic expression vectors--pBV220 and pQE30, and the monomeric form of the recombinant TGF β 1expressed in E.coli DH5α and M15. Sephacryl S 100 HR and Ni + NTA agarose column were used to purify the TGF β 1 protein. With digestion of the enzymes and sequencing, TGF β 1 mature peptide gene was inserted into procaryotic expression vectors--pBV220 and pQE30. After purified, the expressed products of two plasmids in E.coli showed a single protein on SDS--PAGE, and their expression levels were about 15% and 20% of the total bacterial protein. The construction of recombinant plasmids and preparation of the monomeric protein of TGF β 1 lay a solid foundation for further studying the function of TGF β 1 and producing biologically active TGF β 1 protein.
出处
《解放军医学杂志》
CAS
CSCD
北大核心
2002年第5期434-436,共3页
Medical Journal of Chinese People's Liberation Army
基金
国家 973项目资助课题 (编号G1 9990 542 0 4 )
