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目的：通过获得编码血小板生成素（Ｔｐｏ）成熟肽Ｎ端１～１９６个氨基酸的突变体ｃＤＮＡ在大肠杆菌ＪＭ１０９中的表达，为Ｔｐｏ进一步的结构和功能研究提供材料来源。方法：用多聚酶链反应（ＰＣＲ）及ＤＮＡ重组技术，将ＰＣＲ产物克隆到ｐＵＣ１９载体并测序，然后克隆到表达载体ｐＭＡＬ－ｃ２上。结果：转化重组质粒ｐＭＡＬ－ＭＢＰ／ＴｐｏＭ的大肠杆菌经异丙基－β－Ｄ－硫代半乳糖苷诱导４～５小时，ＳＤＳ－ＰＡＧＥ分析显示融合蛋白（ＭＢＰ／ＴｐｏＭ）的分子量约为６．３万。薄层扫描分析表明该蛋白占菌体总蛋白量的３７％。结论：经ＰＣＲ方法扩增的Ｔｐｏ基因突变体可在大肠杆菌中高效表达，为Ｔｐｏ突变体的进一步结构和功能研究以及Ｔｐｏ抗体的制备打下了基础。
OBJECTIVE: To obtain the source of the Tpo for further structural and functional studies by obtaining the cDNA of a mutant cDNA encoding the 1-to 196-amino acids at the N-terminus of thrombopoietin (Tpo) in Escherichia coli JM109. Methods: PCR products were cloned into pUC19 vector and sequenced by polymerase chain reaction (PCR) and DNA recombination techniques, then cloned into the expression vector pMAL-c2. Results: The E. coli transformed with recombinant plasmid pMAL-MBP / TpoM was induced by isopropyl-β-D-thiogalactoside for 4 to 5 hours. SDS-PAGE analysis showed that the molecular weight of the fusion protein (MBP / TpoM) .3 million. Thin layer scanning analysis showed that the protein accounted for 37% of total bacterial protein. Conclusion: The Tpo gene mutant amplified by PCR method can be highly expressed in E. coli, laying a foundation for the further study of the structure and function of Tpo mutant and the preparation of Tpo antibody.