钙结合蛋白D-9K基因原核表达载体的构建、表达产物纯化及其多克隆抗体的制备

THE CONSTRUCTION OF PROKARYOTIC EXPRESSION VECTOR FOR THE GENE ENCODING THE CALCIUM BINDING PROTEIN D-9K, PURIFICATION OF THE EXPRESSED PRODUCT AND PREPARATION OF ITS POLYCLONAL ANTIBODY

本研究构建了pBV220/CabpD-9k-βhCG表达质粒,在大肠杆菌TG-1中进行表达,得到了CabpD-9k-βhCG融合蛋白。SDS-PAGE分析显示融合蛋白的分子量约为34.5KD。West-ern blot结果表明,融合蛋白与兔抗人hCGIgG特异结合。用纯化的融合蛋白免疫C57BL小鼠,制备了多克隆抗体,这为进一步研究钙结 合蛋白D-9k在着床中的功能和子宫内膜Ca^(2+)信号转导奠定了基础。

The expression plasmid pBV220/CabpD-9K-βhCG was constructed,then was transferred into E.Coli TG-1 to express the fusion protein CabpD-9K-phCG. The result of SDS-PAGE showed that CabpD-9K-phCG had an apparent molecular weight of 34. 5KD. Western blot analysis indicated that the product CabpD-9K-|3hCG could be recognized specifically by the rabbit-anti-human hCG IgG. Finally we immunized C57 BL mouse with the purified fused protein to acquire the polyclonal antibody which would lay a foundation of the study on the CabpD-9K function and endometrial calcium signal transduction.