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目的构建IL_3信号肽与内皮抑素融合蛋白基因的真核表达载体。方法用PCR方法扩增IL_3信号肽序列和人内皮抑素基因,然后采用重叠延伸PCR拼接的方法将两者拼接和扩增。并将IL_3-endocl)NA克隆到真核表达载体pcDNA3.1(+)中。结果 IL_3-endo融合蛋白基因包括了IL_3信号肽序列和人内皮抑素蛋白全长基因,基因总长度约701bp,被正确拼接,并克隆到pcDNA3.1(+)的CMV启动子下游。序列分析证实克隆序列、插入方向和读码框架均正确。结论成功的将IL_3信号肽序列连接到人内皮抑素基因序列前并构建了其真核表达载体,为以后的内皮抑素用于基因治疗打下了基础。
Objective To construct the eukaryotic expression vector of IL3 signal peptide and endostatin fusion protein gene. Methods IL - 3 signal peptide sequence and human endostatin gene were amplified by PCR, and then spliced and amplified by overlap extension PCR. IL_3-endocl) NA was cloned into eukaryotic expression vector pcDNA3.1 (+). Results The IL3-endo fusion protein gene contained the full-length IL3 signal peptide sequence and the human endostatin protein. The total length of the gene was about 701bp and was correctly spliced and cloned downstream of the CMV promoter of pcDNA3.1 (+). Sequence analysis confirmed that the cloned sequence, insertion direction and reading frame were correct. Conclusion The IL_3 signal peptide sequence was successfully ligated to the human endostatin gene sequence and its eukaryotic expression vector was constructed, which laid the foundation for future use of endostatin in gene therapy.