sIL-16在大肠杆菌中的表达及活性分析

Expression of sIL-16 in E.coli and Its Activity Analysis

用PCR法扩增本室保存的sIL 16cDNA片段,插入含T7启动子的表达载体pET28a(+)中构建重组质粒pET sIL16,转化大肠杆菌BL21(DE3),低温经IPTG诱导蛋白表达,过Ni2+螯和层析柱一步纯化表达蛋白,重组蛋白与Jurkat细胞共同孵育后,观察它对细胞表面IL 2R表达的影响,结果发现IPTG诱导蛋白表达后,经SDS PAGE电泳,可以看到在18kD左右出现明显蛋白表达条带,表达量占菌体可溶蛋白的40%左右,纯化蛋白的纯度达90%以上,经重组蛋白处理过的Jurkat细胞表面IL 2R的表达水平比未经它处理的细胞增高了18.54%.

The sIL16 cDNA was amplified by PCR,and then the fragment was inserted into an expression vector pET28a(+) under T7 promoter control and constructed recombinant plasmid pETsIL16.pETsIL16 was transformed to E.coli BL21(DE3) and the protein was expressed after IPTG induction at low temperature.It was purified by Ni2+ affinity chromatography column,and incubating with Jurkat cells to access its function to IL2R expression on the surface of the cells. An obvious expression band about 18kD can be detected in the recombinant protein by SDSPAGE.The purity efficiency was up to 90 percent.And the recombinant product upregulated the IL2R expression on surface of the Jurkat cells about 18.54 percent.