目的  对人促红细胞生成素（human erythropoietin，HuEPO）基因进行优化，提高其在大肠杆菌中的表达量以及纯化过程中的回收率。方法  对HuEPO基因的碱基序列进行设计，用大肠杆菌偏爱密码子替换稀有密码子，构建高效表达重组（r）HuEPO（rHuEPO）的基因。设计rHuEPO的氨基酸突变位点，并对这些位点进行定点诱变，获得高亲水性的突变（M）rHuEPO（MrHuEPO）。对rHuEPO和MrHuEPO的三级结构进行预测及对比，验证突变位点设计的合理性。将rHuEPO和MrHuEPO基因克隆至表达载体pET-15b，并在大肠杆菌中进行表达。对表达产物进行初步纯化和复性，比较复性时的rHuEPO和MrHuEPO可溶性和回收率，并用HuEPO活性检测试剂盒检测rHuEPO和MrHuEPO的活性。结果  rHuEPO和MrHuEPO在大肠杆菌中得到成功表达，表达量均＞25%。三级结构预测对比结果表明，突变位点设计合理。rHuEPO和MrHuEPO的表达量无明显差别，但复性时MrHuEPO的可溶性（回收率＞90%）明显高于rHuEPO（回收率<25%）。rHuEPO和MrHuEPO的比活分别为2.36×105 和2.33×105 IU/mg。结论  成功构建了可在大肠杆菌中高效表达rHuEPO和MrHuEPO的基因，并通过定点诱变提高了复性时的MrHuEPO可溶性。

 Objective  Optimize human erythropoietin (HuEPO) gene to improve its expression in E. coli and the recovery in the process of purification. Methods  The base sequence of HuEPO gene was designed by replacing the rare codons in E. coli with biased codons to construct the gene expressing recombinant HuEPO (rHuEPO). The sites of amino acid mutations of HuEPO were designed, and site-directed mutagenesis was performed for these sites to obtain mutational rHuEPO (MrHuEPO) with high hydrophilia. The rHuEPO and MrHuEPO genes of were cloned into expression vector pET-15b and expressed in E. coli, and purification and renaturation of expression products were carried out. Solubility and recovery in the renaturation process were compared between rHuEPO and MrHuEPO. The activities of rHuEPO and MrHuEPO were detected using the HuEPO ELISA kit. Results rHuEPO and MrHuEPO were expressed successfully in E. coli, and the expression levels of both were more than 25%. The results of tertiary structure prediction showed that the design of mutation sites was reasonable. There were no difference in the expression levels between rHuEPO and MrHuEPO, but the solubility of MrHuEPO (recovery ＞90%) was significantly higher than that of rHuEPO (recovery <25%) in the renaturation process. The specific activities of rHuEPO and MrHuEPO were 2.36×105 and 2.33×105 IU/mg,respectively. Conclusion  The genes expressing rHuEPO and MrHuEPO in E. coli are contructed successfully, and the solubility of MrHuEPO in the renaturation process is significantly improved by site-directed mutagenesis.