Expression and purification of TAT/CT-1 and TAT/EGFP fusion proteins / 第三军医大学学报

ABSTRACT

Objective To construct a vector containing protein transduction domain (PTD) and cardiotrophin-1 and a vector containing PTD and EGFP, express them in E. coli. and purify them. To detect the distribution of the two fusion proteins in mice. Methods CT-1 and EGFP were cloned to GST-fusion expression vector pGEX-4T3 by PCR and cloning techniques respectively, and then TAT was cloned into the vectors respectively to give pGEX-TAT/CT-1, pGEX-TAT/EGFP. After induced by IPTG the soluble protein GST-TAT/CT-1 and GST-TAT/EGFP was purified by Glutathione Sepharose 4B. The purified fusion proteins were injected into mice through caudal vein and examined in tissue section by immunohistochemical staining. Results CT-1 and EGFP were effectively amplified and the TAT/CT-1 and TAT/EGFP gene sequencing showed the same sequence as scheduled. The fusion proteins was successfully expressed in E. coli. and purified. Conclusion TAT/CT-1 and TAT/EGFP fusion proteins were expressed and purified successfully. The two fusion proteins were all detected positively in mouse brain, spinal cord, heart and liver.