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PTDSH2融合蛋白的原核表达和纯化
http://www.100md.com 《第四军医大学学报》 2000年第5期
SH2基因;,融合蛋白;表达;纯化;pET16b载体,,SH2基因;,融合蛋白;表达;纯化;pET16b载体,【关键词】SH2基因;融合蛋白;表达;纯化;pET16b载体,0引言,1材料和方法,2结果,3讨论,【参考文献】
     Prokaryotic expression and purification of PTDSH2 fusion protein

    QIU Bo, MA QingJiu, LU JianGuo, YIN JiKai, LI JinMao, WANG Qing

    Department of General Surgery, Tangdu Hospital, Fourth Military Medical University, Xian 710038, China

    【Abstract】 AIM: To construct the recombinant expression vector containing protein transduction domain (PTD) and Src homology 2 (SH2) fusion gene and express PTDSH2 fusion protein in E. coli and purify the fusion protein. METHODS: A 297 bp of human SH2 gene fragment was amplified by PCR and subcloned into pET16b vector downstream of the PTD fragment, an E.coli expression vector, to construct a recombinant plasmid pET16bPTDSH2. The plasmid was transformed into E.coli BL21 (DE3) and induced to express fusion protein PTDSH2 with IPTG. The expression of PTDSH2 was detected by SDSPAGE and Western blot. The expressed protein was purified by NiNTA affinity chromatographic column. RESULTS: SH2 was identical to what reported by GenBank. A novel protein with expected molecular mass (about Mr 15×103) was expressed under the induction with IPTG. The expressed product showed good reactivity to antiHis tag antibody, and was mostly in the form of inclusion bodies. The expressed proteins could be purified via NiNTA affinity chromatography in denatured condition. CONCLUSION: Our successful construction of recombinant express vector pET16bPTDSH2 and efficient expression of PTDSH2 fusion protein in E. coli and purification of interest protein lay a basis for further study on SH2 functions. ......

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