人P选择素胞外区Lectin-EGF片段的基因克隆及表达研究

Gene Cloning and Expression of Human P-Selectin′s Lectin-EGF Domain

目的 构建表达人P -选择素胞外凝集素 -表皮生长因子样区 (Lectin -EGF)片段的基因工程菌。方法 从人血小板中提取总RNA ,经RT -PCR得到P -选择素cDNA ,用PCR方法扩增其中的Lectin -EGF片段 ,并将其克隆到原核表达载体PBV2 2 0中 ,再转化宿主菌DH5α。结果 通过酶切、PCR和基因序列分析确定重组载体片段为目的基因的核苷酸序列 ,并在大肠杆菌中获有效表达重组蛋白。结论 表达人P -选择素胞外区Lectin -EGF片段的基因工程菌的成功构建 。

Objective To construct the genetically engineered bacterium for human P-selectin′s Lectin-EGF domain. Methods Totol RNA was extracted from human platelets, P-selectin cDNA was synthesized by RT-PCR. Lectin-EGF domain gene was amplified with PCR, and cloned into E.coli DH5α. Results The genetically engineered bacterium for human P-selectin′s Lectin-EGF domain was successfully constructed which was verified by restriction enzyme cutting, PCR and sequence analyzing. And a recombinant protein of Lectin-EGF was effectively expressed in E.coli . Conclusion The successful construction of the genetically engineered bacterium for human P-selectin′s Lectin-EGF domain will be providing large quantities of human P-selectin′s Lectin-EGF protein fragments for the making of it′s antibodies.