NT4-Ant-Shepherdin[79-87]融合基因重组载体的构建.docVIP

NT4-Ant-Shepherdin［79-87］融合基因重组载体的构建作者：汤小江，平宝华，潘承恩，杨广笑，王全颖????作者单位：西安交通大学医学院第一附属医院肿瘤外科，陕西西安 710061【摘要】? 目的 设计构建携带NT4-Ant-Shepherdin［79-87］的cDNA融合基因的重组载体，为针对Survivin的靶向治疗奠定基础。方法 应用非对称引物/模板法、PCR技术制备NT4-Ant-Shepherdin［79-87］cDNA片断，连接于pGEM-T-Easy载体，经克隆测序、酶切后与PBV220/NT4质粒连接；转化感受态细胞E.coli DH5α，亚克隆获得NT4-Ant-Shepherdin［79-87］融合基因。结果 克隆出Ant-Shepherdin［79-87］基因，经酶切及测序证实结果正确；连接PBV220/NT4，经克隆、酶切，琼脂糖凝胶电泳证实获得321 bp的NT4-Ant-Shepherdin［79-87］目的基因片断。结论 通过分子生物学技术成功构建了携带NT4-Ant-Shepherdin［79-87］融合基因的重组载体，为进一步研究针对Survivin的靶向抗肿瘤作用奠定了基础 【关键词】? Survivin；Ant；神经营养因子4(NT4)；融合基因；载体 　　Construction of recombinant vector containingfusion gene NT4-Ant-Shepherdin［79-87］ 　　Tang Xiaojiang, Ping Baohua, Pan Cheng’en, Yang Guangxiao, Wang Quanying 　　1. Department of Oncological Surgery, the First Affiliated Hospital, Medical School ofXi’an Jiaotong University, Xi’an 710061;?　　2. Department of Infection Control,the First Affiliated Hospital, Medical School of Xi’an Jiaotong University, Xi’an 710061; 　　3. Department of Geratic Surgery, the First Affiliated Hospital, Medical School of Xi’an JiaotongUniversity, Xi’an 710061; 4. Xi’an Huaguang Bio-engineering Company, Xi’an 710025, China 　　ABSTRACT: Objective? To investigate survivin as an anticancer therapeutic target by use of shepherdin ［79-87］, a novel peptide carrying the survivin sequence from Lys-79 through Leu-87, we constructed an recombinant vector containing fusion gene NT4-Ant-Shepherdin ［79-87］. Methods? The gene of Ant-Shepherdin ［79-87］ was obtained by PCR and T-vector method. After cloned and digested with restricted enzyme, Ant-shepherdin ［79-87］ was inserted in PBV220/NT4 vector. The recombinant vector was transformed into the competent cell, E.coli DH5α. The fusion gene of NT4-Ant-Shepherdin ［79-87］ was identified by agarose gel electrophoresis (AGE). Results? DNA sequencing results verified that the sequence of Ant-Shepherdin ［79-87］ was consistent with what we had designed. After transformed E.coli DH5α, a fragment of 321 bp was c