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目的 :获得具有抑制血管内皮细胞生长活性的重组人内皮抑素蛋白。 方法 :从人胎肝中分离总 RNA,经反转录聚合酶链反应 (RT- PCR)得到 endostatin全基因。用原核细胞表达载体构建了 p BV2 2 0 /endostatin重组质粒 ,经 DNA测序确认后 ,将其转化大肠杆菌 DH5 a进行表达、初步纯化及活性测定。 结果 :经 SDS- PAGE电泳分析表达产物分子量约 2 0 Kda,薄层扫描显示表达产物可达细菌总蛋白的 30 %。 结论 :经生物学活性测定 ,表达蛋白能够抑制 b FGF(碱性成纤维细胞生长因子 )对牛毛细血管内皮细胞 (BCEC)的增殖作用。
Objective: To obtain recombinant human endostatin protein with inhibitory activity on vascular endothelial cells. Methods: Total RNA was isolated from human fetal liver and all the endostatin genes were obtained by reverse transcription-polymerase chain reaction (RT-PCR). The recombinant plasmid p BV220 / endostatin was constructed by using prokaryotic expression vector. After confirmed by DNA sequencing, it was transformed into E. coli DH5a for expression, preliminary purification and activity determination. Results: The molecular weight of the expressed product was about 20 KDa analyzed by SDS-PAGE. The thin layer scanning showed that the expressed product reached 30% of the total bacterial protein. CONCLUSION: The biological activity of the expressed protein can inhibit the proliferation of bovine capillary endothelial cells (BCEC) induced by bFGF (basic fibroblast growth factor).