重组肿瘤导向肽RGD-白喉毒素DT390融合蛋白的原核表达及鉴定

Purpose: To construct the recombinant vector of the fused gene (RGD-DT390) of RGD and truncated diphtheria toxin gene (DT390), express in E.Coli and identify the target protein in vitro. Methods: The fused gene (RGD-DT390) was amplified by PCR, and was inserted into pET28a vector. The recombinant plasmid pET-28a- RGD- DT390 was expressed in E.coli BL21 (DE3) plysS under the induction of IPTG. The fusion protein was identify by SDS-PAGE and Western blot. Results: After induced by IPTG, an extra protein band appeared near the 46KD, more than 20% of the total bacterial protein. Western bloting showed that the recombinant protein could react specifically with a mouse monoclonal antibody against HIS. The expression target accumulates as inclusion bodies. Conclusion: The experiment method provided by this paper can help extract soluble protein RGD- DT390, which will lay a foundation for the research of tumor molecular targeted therapy.