CAJ | 학술논문

目的：通过基因工程技术获得重组结核分枝杆菌19ku蛋白。方法：应用PCR技术扩增卡介苗的19ku蛋白DNA序列；以质粒pET28a为表达载体，构建19ku重组质粒，然后转化大肠埃希菌BL21（DE3）；在异丙基硫代－β—D－半乳糖苷（IPTG）诱导下，分别对不同诱导时间的表达产物进行十二烷基硫酸钠－聚丙烯酰胺凝胶电泳（SDS—PAGE），凝胶经考马斯亮蓝染色检测蛋白。通过镍柱纯化后获得目的蛋白。结果：重组质粒pET28a—p19测序表明与报道的序列相同。它在大肠埃希菌BL21（DE3）细胞内以可溶性形式表达。不同IPTG诱导时间实验表明重组结核分枝杆菌19ku蛋白诱导4h在大肠埃希菌中的表达量最高。结论：pET28a—p19大肠埃希菌工程株可高表达结核分枝杆菌重组19ku蛋白。
목적：통과기인공정기술획득중조결핵분지간균19ku단백。방법：응용PCR기술확증잡개묘적19ku단백DNA서렬；이질립pET28a위표체재체，구건19ku중조질립，연후전화대장애희균BL21（DE3）；재이병기류대－β—D－반유당감（IPTG）유도하，분별대불동유도시간적표체산물진행십이완기류산납－취병희선알응효전영（SDS—PAGE），응효경고마사량람염색검측단백。통과얼주순화후획득목적단백。결과：중조질립pET28a—p19측서표명여보도적서렬상동。타재대장애희균BL21（DE3）세포내이가용성형식표체。불동IPTG유도시간실험표명중조결핵분지간균19ku단백유도4h재대장애희균중적표체량최고。결론：pET28a—p19대장애희균공정주가고표체결핵분지간균중조19ku단백。
Objective: To obtain recombinant 19 ku protein from mycobacterium tuberculosis by using gene engineering technology. Methods: Used PCR technology to amplify 19 ku protein DNA sequence of the BCG vaccine; took plasmid pET28a as expression vector, constructed the 19 ku reeombined plasmid, then transfered E.coli BL21 (DE3) , under the inducement of ispropy-β-D-thiogalactoside (IPTG) , took SDS-PAGE to expression product with different induction time, with coomassie brilliant blue dyeing to detect the protein. The recombinant 19 ku protein was purified by Ni-NTA. Results: The sequencing showed the recombinant plasmid pET28a-p19 had the same sequence with reaport. It expressed in soluble form in DE3. The IPTG induction time experiment showed the recombinant mycobacterium tuberculosis 19 ku protein expressed in E.coli reached highest level under 4 hours inducement. Conclusion : E.coli pET28a-P19 can highly efficiently express the Mycobacterium tuberculosis recombinant 19 ku protein.