CAJ | 학술논문

利用原核表达系统克隆表达斑马鱼p53基因.RT-PCR法从斑马鱼胚胎申扩增获得p53基因编码区,并将其克隆至原核表迭载体pET28a上,构建重组质粒pET28a/z-p53,将重组质粒转化且coli BL21(DE3)受体菌,IPTG诱导表达,表达产物经镍往纯化、尿素透析复性,SDS-PAGE电泳分析,结果表明,p53基因在大肠杆菌中成功表达,表达的p53融合蛋白分子量大约为53kD,透析复性后获得了高纯度可溶性的p53蛋白.
이용원핵표체계통극륭표체반마어p53기인.RT-PCR법종반마어배태신확증획득p53기인편마구,병장기극륭지원핵표질재체pET28a상,구건중조질립pET28a/z-p53,장중조질립전화차coli BL21(DE3)수체균,IPTG유도표체,표체산물경얼왕순화、뇨소투석복성,SDS-PAGE전영분석,결과표명,p53기인재대장간균중성공표체,표체적p53융합단백분자량대약위53kD,투석복성후획득료고순도가용성적p53단백.
The cloning and expressing of zebra fish p53 gene were studied by using prokaryotic expression system. P53 encoding region was obtained from zebra fish embryos by RT-PCR,and then the p53 fragment was cloned in pET28a vector to construct the recombinant plasmid pET28 a/z-p53. It was transformed into E. Coli BL21, and then induced by lmmol/L IPTG to express p53 protein in E.coli BL21 expression system. The recombinant p53 protein was purified by Ni-NTA affinity chromatography under denaturing conditions and renatured through urea gradient dialysis. Soluble p53 protein with high purity was successful obtained.