【摘要】 采用PCR技术 ,根据文献报道的鼠TPO成熟肽基因序列 ,设计并合成两对引物 ,以鼠TPOcDNA为模板 ,扩增获得mT PON端 15 3个氨基酸的 4 78bpcDNA片段及鼠TPO全长 10 32bpcDNA片段 ;mTPO15 3片段与合成的碱性成纤维生长因子序列中Lys119-Lys135aa的 5 1bp肝素结合位点DNA片段连接 ,克隆到M13mp18及M13mp19载体中进行双向测序 ;同时将扩增的鼠TPO全长cDNA片段克隆到M13mp18及M13mp19载体中进行双向测序 ,证明获得鼠血小板生成素与肝素结合位点基因及鼠TPO全长基因 ,继之以pMAL -C2X为表达载体构建表达质粒 ,并经PCR及酶切鉴定更多还原

【Abstract】 Two pairs of primers based on mature peptide of mouse TPO gene sequence was designed according references and PCR amplification was performed,the 478bp cDNA that codes N-terminal of mTPO(153aa) and the 1032bp mTPO cDNA were amplified;mTPO153 DNA fragment was ligated with a 51bp heparin-binding site(Lys119-Lys135aa) DNA fragment in bFGF Then two fragments were cloned into M13mp18 and M13mp19 vector for bidirectional sequencing,the result proved that the cloned genes were mTPO153H and mTPO Both gene were cloned in expressing vector pMAL-C2X,pMAL/mTPO and pMAL/mTPO153H recombinant plasmids were constructed PCR and digest by enzyme were used to test the recombinant更多还原