人PRX3原核表达质粒的构建及表达

Construction of Prokaryotic Expression Plasmid of Human Peroxiredoxin 3 and Its Expression in E.coli

目的构建人Peroxiredoxin 3(PRX3)原核表达质粒,并在大肠杆菌中进行表达。方法采用基因工程技术将PRX3编码序列插入原核表达质粒载体pET-30(a),再将重组质粒转化大肠杆菌BL21(DE3),IPTG诱导表达,并对表达产物进行SDS-PAGE分析。结果酶切鉴定和DNA测序证实人PRX3编码序列全长正确地插入表达质粒中,序列与GenBank报道的一致;重组PRX3在大肠杆菌获得高效表达,且易形成包涵体。结论人PRX3原核表达质粒构建成功,并能在大肠杆菌中高效表达,为后续研究奠定了良好基础。

Objective To construct the prokaryotic expression plasmid of human Peroxiredoxin 3 （PRX3） coding region and express recombinant PRX3 in Escherichia coli. Methods Full length coding region of human PRX3 was inserted into prokaryotic expression plasmid pET-30（a） by gene manipulation techniques. The recombinant plasmid was transformed into E. coli BL21 （DE3） and the expression was induced by IPTG. The products were detected by SDS-PAGE analysis. Results Endonucleases digestion analysis indicated that the coding region of human PRX3 was successfully inserted into the expression vector. DNA sequencing verified that the sequence was identical with that published on GenBank and the reading frame of the recombinant was correct. Recombinant PRX3 was efficiently expressed in E. coli and most of them formed inclusion bodies. Conclusion The prokaryotic expression plasmid of human PRX.3 is successfully constructed and the recombinant is efficiently expressed in E. coll. These laid the foundation for further study on human PRX3.