GST-HPV16E7融合蛋白的原核表达与纯化

目的 构建HPV16地方株E7基因原核表达质粒,使其在原核细胞中高效表达并进行纯化.方法 将成都本地某宫颈癌患者所感染的HPV16 E7全长片段插入原核表达载体pGEX-4T-1,构建重组子pGEX-4T-1-HPV16E7,转化宿主菌E.coli BL-21.经IPTG诱导表达,通过SDS-PAGE、Western blot分析证实蛋白表达的特异性.并用GST亲和层析法对融合蛋白进行纯化.结果 该患者所感染的HPV16为东亚株,成功构建了原核表达载体pGEX-4T-1-HPV16E7并表达出GST-HPV16E7融合蛋白,证实了蛋白表达的特异性,并获得了GST-HPV16E7融合蛋白的纯品.结论 成功表达、纯化了本地流行株GST-HPV16E7融合蛋白,为进一步研究HPV16E7蛋白的功能奠定了实验基础.

Expression and Purification of GST-HPV16E7 Fusion Protein

Objective To construct a recombinant prokaryotic expression vector efficiently expressing HPV16E7 and purify GST-HPV16E7 fusion protein. Methods The HPV16E7 gene obtained from a local cervical cancer patient was cloned into vector Pgex-4T-1, to generate the recombinant named as Pgex-4T-1-HPV16E7 The recombinant plasmid was transformed into E. Coli BL21. After inducing with 1PTG the HPV16E7 fusion protein was analyzed by SDS-PAGE and Western blot. B-PER GST Fusion Protein Purification Kit was appkied to purify GST-HPV16E7 fusion protein. Results The recombinant plasmid Pgex-4T-1-HPV16E7 was successfully constructed. Highly expressed and purified GST-HPV16E7 fusion protein was obtained. The specificity of fusion protein was verified by SDS PAGE and Western blot. Conclusion GST-HPV16E7 fusion protein was successfully expressed and purified.