| 曹华,葛忠良,张群伟,等.rhTPO的克隆及其在原核细胞中的表达[J].中华放射医学与防护杂志,1997,17(5):316-318.Cao Hua,Ge Zhongliang,Zhang Qunwei,et al.Cloning and prokaryotic expression of rhTPO in prokaryotic cells[J].Chin J Radiol Med Prot,1997,17(5):316-318 |
| rhTPO的克隆及其在原核细胞中的表达 |
| Cloning and prokaryotic expression of rhTPO in prokaryotic cells |
| 投稿时间：1995-12-22 修订日期：1996-04-29 |
| DOI： |
| 中文关键词: 血小板生长因子 克隆 原核表达 |
| 英文关键词:Thrombopoietin Prokaryotic expression Cloning |
| 基金项目: |
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| 中文摘要: |
| 目的 为了更加简便有效地得到重组人血小板生长因子(rhTPO),以治疗放射及化学药物等所致骨髓损伤造成的血小板减少与缺乏,缓解临床上的出血症状,降低死亡率。方法 本实验从6月龄人胚胎肝脏细胞中分离mRNA,设计合成特异性引物,经逆转录,套式PCR等步骤克隆出了人TPO全基因。将人TPO基因亚克隆至原核细胞表达载体pBV220,形成了重组表达质粒pBV220/TPO,以该重组质粒转化了原核细胞大肠杆菌DH5α,应用42℃热诱导表达目的蛋白。结果 经SDS-聚丙烯酰胺凝胶电泳检测,得到了TPO在原核细胞中的表达,其分子量与目的一致。结论 hTPO可以在原核细胞得到良好的表达,为临床血小板减少症的生物治疗奠定了基础。 |
| 英文摘要: |
| Objectives Thrombopoietin (TPO) is a normal physiological hematopoietic factor which is able to stimulate the hematopoietic progenitor cells to proliferate and differentiate into megakaryocytes and to induce the production and release of platelets. In this study rhTPO was cloned to be potentially used for treatment of radiation-induced thrombocytopenia, we studied the prokaryotic expression of rhTPO.Methods According to the cDNA sequence encoding hTPO published elsewhere, we designed two sets of PCR primers at first, and cloned hTPO gene from fetal liver mRNA by means of RT PCR, then subcloned it into prokaryotic expression vector pBV220 to form recombinant expression plasmids pBV220/TPO, and to transfer it into prokaryotic cells escherichia coli, DH5α. Expression was induced by heat of 42℃. Results We obtained the aimed prokaryotic expression of recombinant(r) hTPO by assessment of SDS-Page. Conclusion hTPO can be expressed completely in prokaryotic cells. The prokaryotic expression of rhTPO might be a basis for the bio treatment of radiation induced thrombocytopenia. |
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