Study On Construction And Expression Of Common IFN And THY α1 Fusion Gene

In the present study, expression and purification of recombinant thymic peptide and IFN were reported. The combinated IFN and THYal is more powerful than the individual of both IFN and THYalmedicines during treating human cancers and some virus infection diseases, and we considered to construct a fusion protein molecule containing both IFN and THYal by DNA recombination. First of all, based on the three dimension structure of both molecules, a linker with 14 amino acid length was designed to connect IFN N terminal and THYal C terminal. After that, the chemically synthesized cDNA of thymic peptide al was amplified by PCR, then splicing the IFN gene by overlapping PCR. and inserted in to PGEM-T vector. Then there combinant plasmid was transformed into DH5a cells, After duplication, The positive recombinant plasmid was identified by double enzyme digestion and PCR and then the sequence was analyzed by computer. The result showed that the gene fragment amplified was 647 bp long and the homology with IFN encoding sequence from Genebank was 70%. It is confirmed that the gene fragment obtained was the very common IFN gene。The expression of IFNTHY gene in prokaryotic cells and determination of its biological activities, the thymic peptide al and IFN fusion gene fragment was inserted into PET28a to construct thymic peptide al IFN fusion gene and transformed into host cells, BL21(DE3)and induced by IPTG.. The specific protein expressed (about 22.8 kDa) was detected by SDS-PAGE and Westerblot. Then the fusion proteinwas purified by Zn-Sepharose affinity chromatography. The yield of fusion protein was 35 mg from 1 liter bacterial culture medium affinity chromatography of Zn-Sepharose 3.8 mg of thymic peptide al.