耐辐射奇球菌超氧化物歧化酶基因的克隆和表达

目的 构建耐辐射奇球菌(D．rdiodurans)锰超氧化物歧化酶(Mn-SOD)基因的原核表达重组子，并在E．coli BL21(DE3)中表达。方法 用PCR方法自D.radiodurans基因组中扩增Mn-SOD目的片段。将该基因与原核表达质粒载体pET-30a( )连接，构建重组质粒pET-SOD，并转化表达宿主菌E．coli BL21(DE3)。用异丙基硫代-β-D-半乳糖苷(IPTG)诱导重组SOD蛋白表达，SDS-PAGE分析表达产物。结果 获得了D．radiodurans-Mn-SOD基因的pET原核表达重组质粒，该质粒经IPTG诱导能在E．coli BL21(DE3)中高效表达目的蛋白，蛋白活性可达51800u／g湿菌体。结论 成功构建了原核表达重组质粒pET-SOD，实现了SOD在原核细胞中的高效表达，表达产物活性较高，为重组D．radiodurans Mn-SOD的进一步研究和应用奠定了基础。

Cloning and Expression of Superoxide Dismutase Gene from Deinococcus radiodurans in E. coli

OBJECTIVE: To construct expressing recombinant of Mn-SOD of Deinococcus radiodurans and express the target protein in E. coli BL21(DE3). METHODS: SOD gene was amplified by PCR from genomic DNA of Deinococcus radiodurans and inserted into expression plasmid pET-30a(+) to create the recombinant pET-SOD. After being analyzed by the restriction endonuclease, the plasmid was transformed into E. coli BL21(DE3), and the recombinant protein was expressed after induction by the isopropyl-beta-D-thiogalactopyranoside (IPTG) and was analyzed with SDS-PAGE. RESULTS: The recombinant plasmid pET-SOD was obtained, and the recombinant protein was highly expressed in E. coli BL21(DE3). The activity of recombinant superoxide dismutase was 51,800 U per gram of wet bacteria. CONCLUSION: This study has provided a foundation for further studies and applications of the recombinant Mn-SOD.