超抗原VEGF-SEA融合基因的克隆与表达  

Cloning and Expression of Superantigen Fusion Genes VEGF-SEA

作　　者：边小鹤[1] 孙嘉琳[1] 朱宏丽[1] 罗金凤[1] 胡坤[1] 

机构地区：[1]天津科技大学生物工程学院,300457

出　　处：《天津医药》2008年第10期783-785,共3页Tianjin Medical Journal

基　　金：天津市科委应用基础研究重点项目(项目编号:06YFJZJC02600)

摘　　要：目的:构建血管内皮生长因子-葡萄球菌肠毒素A(VEGF-SEA)基因的原核表达质粒,获得VEGF-SEA融合蛋白。方法:制备VEGF-SEA基因片段,插入pET40b构建重组质粒pET40b-VEGF-SEA,经序列分析正确后转化大肠杆菌BL21(DE3),进行IPTG诱导表达蛋白。用十二烷基磺酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)分析表达蛋白的相对分子质量及表达形式,并对产物采用His·Bind Buffer kit试剂盒纯化。结果:获得VEGF-SEA基因片段长1130bp,经序列分析与预期完全一致,其表达出目的蛋白的大小约为66.2ku,以包涵体形式表达,经纯化,纯度达到90%以上。结论:表达载体pET40b-VEGF-SEA构建成功,VEGF-SEA融合蛋白获得高效表达,本实验为进一步研究VEGF-SEA蛋白的活性及其功能,探讨超抗原抑制肿瘤生长作用奠定了基础。Objective： To construct prokaryotic expression plasmid of VEGF-SEA gene, and to get the fusion proteins. Methods： VEGF-SEA gene was synthesized, and was cloned into pET40b to construct recombinant plasmid pET40b-VEGF-SEA. The gene was inserted into BL21 （DE3） of E.coli after sequencing for inducing the expression recombinant proteins. The relative moleculer size and form of the proteins were analyzed by SDS-PAGE, and the products were purified by His· Bind Buffer kit. Results： The length of PCR product was 1 130 bp and identical with what the GenBank reported after sequencing. The relative moleculer size of the expressed proteins were 66.2 ku, and were in the forms of including bodies. The purity of target protein was more than 90%. Conclusion： The prokaryotic expression plasmid was constructed successfully, and was induced to express high level recombinant proteins. The expression of recombinant VEGF-SEA protein with natural activity lays a basis for further study of VEGF-SEA proteins＇ function in controlling tumors.

关 键 词：血管内皮生长因子类 葡萄球菌属 肠毒素类 超抗原 基因表达 克隆 分子 

分 类 号：R735.7[医药卫生—肿瘤] R733.7[医药卫生—临床医学]