人血小板生成素在重组痘苗病毒中的分泌表达

Secreting expression of human thrombopoietin in recombinant vaccinia viruses

目的研究人血小板生成素（ｈＴＰＯ）在重组痘苗病毒中的表达。方法以ＰＣＲ法从人胚肝ｃＤＮＡ文库中克隆完整的ｈＴＰＯ基因，将所克隆基因插入痘苗病毒表达质粒ｐＪＳＡ１１７５ＳｍａⅠ位点，置于７．５Ｋ启动子控制之下，并构建在３４９位氨基酸变异与非变异的两种ＴＰＯ重组痘苗病毒。以Ｄｏｔ－ＥＬＩＳＡ法和促ＴＰＯ依赖细胞增殖的活性测定，进行表达产物的检测。结果研究证实：ｈＴＰＯ可分泌性的表达于重组痘苗病毒感染细胞上清中，并具有特异性的生物学活性。结论ｈＴＰＯ在重组痘苗病毒中的表达为ＴＰＯ结构与功能研究提供了一条新途径。

Objective To study the expression of human thrombopoietin (hTPO) in recombinant vaccinia viruses. Methods The intact hTPO gene was cloned from a human embryo liver cDNA library with PCR. Two of cloned genes were inserted at Sma Ⅰ site of pJSA1175 expression vector under the control of 7 5K promoter and the recombinant plasmids were subsequently used to construct recombinant vaccinia viruses. The rhTPO expressed from vaccinia recombinants were detected with Dot ELISA and the bioassay using an engineered TPO dependent cell line. Results It was confirmed by immunoassay and bioassay that rhTPO expressed in vaccinia recombinants infected cells could be secreted into the culture medium with specific biological activity. Conclusion This work has provided a new approach to study the structure and function of hTPO.