新型噬菌体展示载体pCANTAB5S的构建  

Construction of a novel phagemid pCANTAB5S

作　　者：徐容[1] 潘卫[2] 沈毅 陈秋莉[2] 潘欣[2] 冯春芝[2] 戚中田[2] 刘彦君 邓松华[1] 

机构地区：[1]安徽医科大学病理生理学教研室,合肥230032 [2]第二军医大学微生物学教研室,上海200433 [3]复旦张江生物医药股份公司,上海200000

出　　处：《安徽医科大学学报》2004年第2期83-86,共4页Acta Universitatis Medicinalis Anhui

基　　金：国家" 863"计划资助项目 (编号 :2 0 0 1AA2 15 0 5 1;2 0 0 2AA2Z3 3 0 9);国家自然科学基金资助项目 (编号 :3 0 170 85 6)

摘　　要：目的　进一步改造噬菌体展示载体 ,使之能更有效地展示各种外源随机多肽文库。方法　以 pCANTAB5X IFN α2b重组质粒为模板 ,在上游引物中引入XbaⅠ、SacⅠ酶切位点 ,在下游引物中引入SacⅠ、StuⅠ、SalⅠ酶切位点 ,将用该引物扩增的IFN α2b衔接片段PCR产物克隆到pMD18 T中 ,再将该片段用XbaⅠ和SalⅠ酶切并将之插入pCANTAB5L的XbaⅠ和SalⅠ位点中 ,经SacⅠ酶切 ,线性载体片段自连 ,构成新型噬菌体展示载体 pCANTAB5S。 结果　限制酶切位点SacⅠ被引入到噬菌体展示载体 ,并校正了pCANTAB5L载体StuⅠ、SalⅠ等位点的读框。结论　成功构建了含读框正确的SacⅠ等多个克隆位点的新型噬菌体展示载体pCANTAB5S ,可用于各种外源随机多肽文库的有效展示。Objective To further reconstruct the new phage display vector for more effectively displaying the foreign random peptide library. Methods The recombinant plasmid pCANTAB5X-IFN-α2b was provided as the amplification template,new cloning site Sac Ⅰ was added to upstream primer,and Sac Ⅰ,Stu Ⅰ and Sal Ⅰ were added to downstream primer for linker IFN-α2b PCR amplification. The PCR product was cloned into pMD18-T. This DNA fragment was cut out by disgestion of restriction enzymes Xba Ⅰ and Sal Ⅰ,and inserted into the clone sites in phagemid pCANTAB5L,by digestion of restriction enzyme Sac Ⅰ,the linear vector fragment was isolated and ligated to construct the new phage display vector pCANTAB5S. Results A new cloning site of Sac Ⅰ was added to phagemid pCANTAB5S and the open reading frame of the clone sites Stu Ⅰ,Sal Ⅰ,Kpn Ⅰ etc. In phagemid pCANTAB5L was corrected. Conclusion The novel phagemid with a Sac Ⅰ clone site and a correct open reading frame of multi-clone sites is successfully constructed which can effectively display the foreign random peptide library.

关 键 词：噬菌体 pCANTAB5S 遗传学 质粒 遗传载体 肽库 

分 类 号：R373.9[医药卫生—病原生物学] R394[医药卫生—基础医学]