人酸性成纤维细胞生长因子的高效表达

构建能高效表达人酸性成纤维细胞生长因子（haFGF)的基因工程菌，优化产物分离纯化条件，并对纯化产物进行理化性质鉴定，采用PCR方法扩增haFGF cDNA，将该cDNA片段插入表达载体pET3c的表达框架中，筛选得到了重组质粒pET3c-haFGF，转化大肠杆菌BL21（DE3），构建了表达菌件BL21（DE3）／pET3c-haFGF.IPTG诱导表达，通过离子交换，肝素亲和两步层析的方法分离纯化产物，用MTT法测定产物的活性，haFGF的表达量约为20％，经过两步层析纯化，获得了电泳纯的haFGF样品，纯化haFGF样品的比活性为ED50小于4.0ng/mL.BL21(DE3)/pET3c表达系统能高效表达haFGF,经过纯化得到了理化性质及生物活性与天然haFGF对照品一致的重组人酸性成纤维细胞生长因子。

Construction of High-Level Expression Strain of Human Acidic Fibroblast Growth Factor

High_level expression system of human acidic fibroblast growth factor(haFGF) was constructed. For this purpose, haFGF cDNA produced by PCR was cloned into the expression vector pET3c. The expression was induced by IPTG. The expressed haFGF was purified by ionic exchange and heparin affinity chromatography from the supernant of bacteria lysate. The results showed that the expression level of haFGF in BL21(DE3)/pET3c_haFGF was about 20%. The specific activity of the purified haFGF was less than 4.0 ng/mL. Thus, it was concluded that haFGF was expressed in high level by BL21(DE3)/pET3c expression system. The characters of the recombinant haFGF were identical to those of natural haFGF.