重组人类抗砷相关基因在大肠埃希菌的表达

Expression of recombinant hARRG cDNA in E.coli and purification of hARRG protein

目的 构建含重组人类抗砷相关基因 (humanarsenicresistencerelatedgene ,hARRG)的表达载体 ,诱导其在转化菌表达 ,分离纯化表达蛋白 ,研究该蛋白质的理化性质、抗砷功能和免疫活性 ,深入研究人类对砷化物的抵抗作用。方法 将hARRGcDNA开放阅读框亚克隆到原核表达载体Pet11C中 ,用异丙基 - β -D -硫代半乳糖苷 (IPTG)诱导表达蛋白质 ,利用阴离子交换柱Sepharose纯化蛋白质 ,SDS -PAGE胶电泳观察结果。 结果 将hARRGcDNA成功亚克隆到原核表达载体Pet11C中 ,并成功在大肠杆菌中表达 ,表达的hARRG蛋白占菌体蛋白的 5 %左右 ,该蛋白质被分离纯化。结论 原核表达载体Pet11C可以在大肠杆菌中表达hARRGcDNA 。

Objective To construct expression vector of the recombinant human arsenic resistance related gene (hARRG),induce its expression in DE\-3 and isolate and purify expression product,for studying the physiochemistry characteristic,function and immune activity of the protein,and further researching the arsenic resistant effects of human.Methods hARRG cDNA was subcloned into prokaryotic expression vector Pet11C.The recombinant protein expression was induced by IPTG,then,the protein was purified by anions Ion-exchange column Sepharose and examined by SDS-PAGE gel.Results hARRG cDNA was successfully subcloned into prokaryotic expression vector Pet11C and expressed in E.coli and the protein was purified by anions Ion-exchange column successfully.Conclusion Pet11C excpression vector containing hARRG cDNA wassuccessfully constructed,the cell DE\-3 transformed with expression vector capable of expression the gene and a hARRG protein could be purified by anions Ion-exchange column Sepharose.