文档介绍：1 重组人血管内皮生长因子 D 诱导鸡胚尿囊膜血管增生作者:陈浩丁秀云高媛柳晓兰高建恩孙启鸿【摘要】为了表达和纯化具有生物学活性的人血管内皮生长因子(vascular endothelial growth factor , VEGF) 拟D, 探讨 VEGF 拟D 对血管增生作用,从人胎肺组织中提取总 RNA , 设计特异性引物并应用 RT 拟 PCR 法扩增 VEGF 拟D 成熟肽片段; 经测序证明目的片段序列正确后, 构建重组表达质粒 pGEX 拟 5X 拟 1/VEGF 拟D 并在大肠杆菌 BL21 ( DE3 ) 中进行 GST 拟 VEGF 拟D 融合蛋白的表达。结果表明,融合蛋白的分子量约为 38 kD , 表达量占菌体总蛋白的 15% 以上, 抗 GST 和抗 VEGF 拟D 的抗体均能识别融合蛋白;纯化的 GST 拟 VEGF 拟D 融合蛋白与 VEGFR 拟 3/Fc 具有结合活性, 并能刺激红白血病细胞系 HEL 细胞生长; 在鸡胚尿囊膜实验中 GST 拟 VEGF 拟D 具有促进鸡胚尿囊膜血管生成的作用。结论:成功表达了具有生物学活性的重组人 VEGF 拟D 融合蛋白,为进一步研究 VEGF 拟D 的作用机理提供了实验模型。 2 【关键词】血管内皮生长因子 D; 血管内皮生长因子受体 3 ;鸡胚尿囊膜;血管生成 binant Human VEGF 拟D Induces the Angiogenesis of the Chick Embryo Chorioallantoic Membrane Abstract Vascular endothelial growth factor 拟D (VEGF 拟 D) and vascular endothelial growth factor receptor 拟 2,拟3 (VEGFR 拟 2,拟 3) with thEir corresponding signaling pathway play significant roles in the development of the embryonic vascular system and pathological lymphangiogenesis. The study was aimed to express and purify the GST 拟 VEGF 拟D fusion protEIn, and to explore the angiogenesis effect of VEGF 拟 D. The total RNA was extracted from human fetal lung tissue, and the mature form of VEGF 拟D was expanded by polymerase chain reaction (PCR), then the plasmid pGEX 拟 5X 拟 1/VEGF 拟D was reconstructed and the GST 拟 VEGF 拟D fusion protein expressed in transformed BL21 拟 results showed that the molecular mass of this fusion protein was 38 kD passed more than 15% of the total bacteria fusion protein was recognized by anti 拟 GST and anti 拟 VEGF 拟 3 D soluble GST 拟 VEGF 拟D fusion protein could interact with VEGFR 拟 3/Fc and was able to stimulate the proliferation of human erythroleukemia cell line ( HEL ) cells. The data of chick embryo chorioallantoic membrane (CAM) experiments indicated that GST 拟 VEGF 拟D could induce the CAM angiogenesis. It is concluded that the GST 拟 VEGF 拟D fusion protein with biological activity was essfully expressed, and which may provide an experiment