| 生长抑素噬菌体展示鼠源单链抗体库的构建与鉴定 |
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| 引用本文： | 张颖,侯化鹏,王海瑞,白莉雅,杨利国.生长抑素噬菌体展示鼠源单链抗体库的构建与鉴定[J].农业生物技术学报,2009,17(3):375-380. |
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| 作者姓名： | 张颖 侯化鹏 王海瑞 白莉雅 杨利国 |
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| 作者单位： | 1. 华中农业大学动物科技学院动物遗传育种与繁殖教育部重点实验室,武汉,430070
2. 华中农业大学动物医学院,武汉,430070 |
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| 基金项目： | 国家自然科学基金，国家支撑计划项目 |
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| 摘 要： | 应用噬菌体表面展示技术，从生长抑素免疫的小鼠脾脏中提取总RNA，用RT-PCR技术反转录成cDNA，通过PCR扩增出抗体重链可变区VH基因和轻链可变区VL基因，再用编码(G1y4Ser)3的Linker在体外将VH和VL连接成单链抗体(ScFv)基因，克降到噬菌粒载体pCANTAB5E中，电转化至感受态的大肠杆菌TG1,经辅助噬菌体M13K07超感染，形成噬菌体单链抗体库，有效库容为9.3×107。为利用亲和富集筛选技术，获得具有与SS结合活性的完整重组噬菌粒克隆奠定了基础。
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| 关 键 词： | 生长抑素 噬菌体展示 单链抗体 |
| 收稿时间： | 2008-3-10 |
| 修稿时间： | 2008-4-28 |
Construction of a Phage Display Library of Repertoire Single Chain Antibody from Mouse Immunized against Somatostatin and Its Identification |
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| ZHANG Ying,HOU Hua-peng,WANG Hai-rui,BAI Li-ya,YANG Li-guo.Construction of a Phage Display Library of Repertoire Single Chain Antibody from Mouse Immunized against Somatostatin and Its Identification[J].Journal of Agricultural Biotechnology,2009,17(3):375-380. |
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| Authors: | ZHANG Ying HOU Hua-peng WANG Hai-rui BAI Li-ya YANG Li-guo |
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| Abstract: | Using phage display technique we constructed a library of repertoire immunoglobulin from mouse immunized with SS. The heavy-chain and light－chain variable region gene (VH and VL) repertoire of immunoglobulin were amplified individually from the spleen cell RNA by RT－PCR and joined by a DNA linker encoding (G1y4Ser)3 as a single chain (ScFv) DNA fragment with overlap extension. These fragments were cloned into the phagemid pCANTAB5E and the recombinant phagemid was transformed to susceptible E.coli TGl by electroporation.In the presence of helper phage M13KO7, the ScFv fusion protein were displayed on the surface of recombinant phages. The phage antibody library was generated and its size was determined to be 9.3×107,these results are bases to further phase antibody library construction |
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