基因工程技术获取重组人血小板因子4

目的：用基因工程方法获取重组人血小板因子4（rhPF4），为下一步基础理论研究和临床应用奠定基础。方法：PCR方法获得人PF4编码序列，克隆入质粒pRSET，构建融合表达载体pRSET—PF4，转化大肠杆菌，以IPTG诱导表达融合的人PF4蛋白，经镍柱亲和层析纯化，SDS—PAGE分析所表达的目的蛋白及纯化后蛋白。结果：成功构建了表达载体pRSET—PF4，DNA序列测定结果与预期结果一致。IPTG诱导表达的rh—PF4融合蛋白部分以可溶形式存在，占菌体总蛋白量的11％。亲和层析纯化后目的蛋白纯度为84％。结论：获得原核基因工程表达的rhPF4蛋白。

Obtaining of human platelet factor 4 by means of genetic engineering technique

Objective： To obtain human platelet factor 4 （hPF4） by means of genetic engineering technique. Methods：DNA coding sequence for hPF4 was obtained by PCR and cloned into plasmid pRSET to construct fusion expressed vector pRSET- PF4. After transformed pRSET -PF4 to E. coli DH5α, the bacteria were induced by IPTG. The expressed hPF4 fused protein was purified by Ni - NTA affinity chromatography and identified by SDS - PAGE. Results ： The DNA sequencing showed that the expression vector pRSET - PF4 was constructed successfully. After induced by IPTG, the target protein accounted for 11% of the total bacterial protein and partly was in a soluble form. The purity of the fused protein was up to 84% after Ni - NTA affinity chromatography. Conclusion：The recombinant hPF4 fused protein is obtained.