CAJ | 학술논문

目的：表达、纯化T7噬菌体尾蛋白P11并进行鉴定。方法：PCR扩增T7噬菌体尾蛋白P11基因片段， EcoRⅠ和XhoⅠ双酶切后连接表达载体pTIG-TRX，重组表达载体转化大肠杆菌BL21（DE3），经IPTG诱导表达，纯化目的蛋白，SDS-PAGE分析P11蛋白的相对分子质量大小，并多次免疫新西兰大白兔制备其多克隆抗体，Western blot鉴定蛋白活性。结果：PCR扩增P11基因后，成功诱导表达了含6×His标签的P11蛋白，SDS-PAGE显示所表达蛋白的相对分子质量约为25000，通过多次免疫获得了抗P11的多克隆抗体并对其纯化；制备的多克隆抗体能够识别T7噬菌体和P11蛋白，具有较高特异性。结论：成功表达了P11蛋白并制备了其多克隆抗体，为T7噬菌体展示系统的应用奠定了基础。
목적：표체、순화T7서균체미단백P11병진행감정。방법：PCR확증T7서균체미단백P11기인편단， EcoRⅠ화XhoⅠ쌍매절후련접표체재체pTIG-TRX，중조표체재체전화대장간균BL21（DE3），경IPTG유도표체，순화목적단백，SDS-PAGE분석P11단백적상대분자질량대소，병다차면역신서란대백토제비기다극륭항체，Western blot감정단백활성。결과：PCR확증P11기인후，성공유도표체료함6×His표첨적P11단백，SDS-PAGE현시소표체단백적상대분자질량약위25000，통과다차면역획득료항P11적다극륭항체병대기순화；제비적다극륭항체능구식별T7서균체화P11단백，구유교고특이성。결론：성공표체료P11단백병제비료기다극륭항체，위T7서균체전시계통적응용전정료기출。
To express,purify and identify capsid tail protein P11 of T7 bacteriophage.Methods:Gene of pro-tein P11 was amplified by PCR , digested with restriction endonuclease EcoRⅠand XhoⅠand then inserted into the expres-sion vector pTIG-TRX.The recombinant vector was transformed to E.coli BL21(DE3).After IPTG induction,the recombi-nant protein was purified by metal chelate chromatography .Its molecular weight was identified by SDS-PAGE and its activity was analysed by Western blot .The recombinant protein P 11 was used to immune rabbit .Finally,the rabbit polycolonal anti-body against protein P11 were obtained.Results:Gene of protein P11 was amplified by PCR.The recombinant protein P11 with 6 ×His tag has been successfully expressed after induction .SDS-PAGE analysis suggested that the molecular weight of the expressed protein was approximately 25 000 .Western blot analysis suggested that the polycolonal antibodidy could rec-ognize T7 phage and P11.Conclusion:We have purified P11 fusion protein and prepared the rabbit polycolonal antibody a-gainst protein P11 successfully ,which lays foundation for the application of T 7 phage display .