果树学报2013，Vol.30Issue(2)：185-190,6.
桃果实PpCuZnSOD基因的原核表达、纯化与鉴定
Prokaryotic expression, purification and characterization of PpCuZnSOD gene of Prunus persica
摘要
Abstract
[Objective]This stady was to achieve stable and high efficient expression of peach copper-zinc superoxide dismutase (CuZnSOD) in the Escherichia coli and obtain engineering bacterium with high purity and strong activity. [ Method]PpCuZnSOD gene (GenBank No. JX217743) was inserted into the vector pET-32a ( + ) to construct fusion vector pET-32a ( + )-PpCuZnSOD, which was transformed into E. coli BL21 (DE3) cell and then was induced by IPTG. The recombinant protein was purified with nickel affinity chromatography column, detected with SDS-PAGE and Western blotting. The concentration of the recombinant protein was determined by Brad Ford method, and the enzymatic activity was determined by xan-thine oxidation method.[Result]The prokaryotic expression vector of pET-32a(+)-PpCuZnSOD was constructed successfully, and the molecular weight of fusion protein induced by IPTG was about 35 kDa. The concentration of the recombinant protein was 183.7 mg·L-1,and the enzyme activity was 5.23×103 U·mg-1. [Conclusion]The stably and highly expressed prokaryotic expression vector of pET-32a( + )-PpCuZnSOD was constructed successfully, and the recombinant protein had high biological activity. This study laid the foundation for further study of PpCuZnSOD.关键词
桃/PpCuZnSOD/原核表达/纯化/鉴定Key words
Prunus persica L., PpCuZnSOD/ Prokaryotic expression/ Purification/ Characterization分类
农业科技引用本文复制引用
吴军帅,丛丽君,段艳欣,董晓颖,王滨,李培环..桃果实PpCuZnSOD基因的原核表达、纯化与鉴定[J].果树学报,2013,30(2):185-190,6.基金项目
国家自然科学基金青年科学基金(NSFC,30900976) （NSFC,30900976）
山东省高等学校科技计划(J12LF06) （J12LF06）
