Study On Expression Of A Synthesized Gene For Human Epidermal Growth Factor In E.coli And Bombyx Mori And Application

A 168bp gene sequence (HChEGF) encoding 53 amino acid residues of human epidermal growth factor (hEGF) was synthesized. In view of the preference of codon for silkworm and E. coli, the synthesized HChEGF nucleotide acid is different to that of natural hEGF. The sequence was cloned into pET-28a(+) between the sites of BamH I and EcoR I . The HChEGF expression was performed in E.coli. SDS-PAGE and Western blot suggests that the recombinant HChEGF has the molecular weight of about l0 kDa and is expressed in a high expression level of above 30% of the total protein. Through the single-step metal chelate affinity chromatography, the recombinant HChEGF showed an almost homogenous protein band on SDS-PAGE. The HChEGF was cloned into pBacPAK-His(l) to form pBacPAK-His-HChEGF plasmid. Recombinant virus Bm-Bac-HChEGF was generated by cotransfection into Bm-N cells with the transfer plasmid pBacPAK-His-HChEGF and the modified Bombyx mori nuclear polyhedrosis virus Bm-BacPAK6 genomic DNA. SDS-PAGE shows that recombinant protein expressed in pupa is about lOkDa in molecular weight. And the expression level was 33.07 ng per milliliter hemolymph detected by ELISA. The hEGF sample expressed in B. mori improved the growth of kidney cells of SD rats. And after taking hEGF sample expressed in B. mori orally, higher level of hEGF could be detected from serum of SD rats. And the expression product also shows effects to gastric mucosa lesion induced by acetic acid.