摘要
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。
AIM To construct the eukaryotic expression vector which expresses the fusion protein of IL_3 signal peptide and human endostatin (endo)protein. METHODS IL_3 signal sequence and human endostatin gene were amplified with PCR and spliced with overlap extension PCR. The IL_3-endo cDNA was cloned into mammalian expression vector pcDNA3.1 (+). RESULTS The chimeric gene, about 701bp, which included IL_3 signal sequence and entire human endostatin gene, was successfully spliced and cloned into eukaryotic expression vector pcDNA3.1 (+) downstream from the CMV promoter. The data of nucleotide sequence determination showed that the chimeric cDNA sequence was the same as reported sequence, and in one ORF. CONCLUSION The recombinant vector containing chimeric gene of IL_3-endostatin can express IL_3-endostatin protein in mammalian cell and secrete to extracellular matrix from cells. The success of constructing this vector laid the foundation for studying tumor gene therapy by endostatin.
出处
《世界华人消化杂志》
CAS
2001年第1期43-46,共4页
World Chinese Journal of Digestology
基金
国家自然科学基金
No.39970819
关键词
信号肽类
内皮抑素
基因疗法
真核表达载体
聚合酶链反应
基因扩增
signal peptides
endostatin
gene therapy
eukaryotic expression vector
polymerase chain reaction
gene amplification
