小鼠cbfa1部分编码区在大肠杆菌中的表达和纯化

The expression and purification of mouse cbfal in E.coli

目的 在大肠杆菌中表达cbfa1肽段并进行纯化 ,为进一步制备抗体奠定基础。方法 构建 pRSETA -cbfa1原核表达重组质粒 ,转化大肠杆菌BL2 1,IPTG诱导蛋白表达 ,采用Ni-NTA亲和层析柱进行初步纯化。结果 成功地将 2 0 4bp的cbfa1片段插入 pRSETA原核表达载体中 ,且位于T7启动子下游、读框正确 ,在0 .1mmol/L的IPTG诱导下 ,SDS -PAGE电泳可见在约 14KD处有蛋白新生带 ,用Ni-NTA柱亲和层析纯化融合蛋白 ,得到的cbfa1蛋白纯度大于 95 %。

Objective To express and gain cbfa1 fusion proteins in E.coli. Methods A 204bp segment comprised of part of cbfa1 runt domain and nucleus location signal was selected and it was cloned to express vector pRSET A.The recombinant plasmid was transformed into E.coli BL21 and induced by IPTG to express fusion proteins.The recombinant proteins were purified by metal-chelate affinity chromatography on Ni-NTA resin.Results SDS-PAGE showed that a new protein band about 14KD was gained and purified. Conclusion The fusion proteins encoded by the transcription factor cbfa1 could be gained in E.coli induced by IPTG in vitro.