Expression of human granulocyte‐macrophage colony‐stimulating factor gene in insect cells by a baculovirus vector

FEBS Letters

Chiou, Chuang-Jiun¹  Wu, Ming-chi¹ 

[1] Department of Biochemistry, University of North Texas/Texas College of Osteopathic Medicine, Denton, TX76203, USA

关键词: Colony-stimulating factor;    Baculovirus;    Gene expression;    (Spodopterafrugiperda);    hGM-CSF;    human granulocyte-macrophage colony-stimulating factor;    Sf9;    Spodoptera frugiperda;    AcNPV;    Autographa californica nuclear polyhedrosis virus;    SDS-PAGE;    sodium dodecyl sulfate-polyacrylamide gel;    hpi;    hour postinfection;   

DOI  :  10.1016/0014-5793(90)80020-J

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A plasmid pAc373GM-CSF was constructed and co-transfected into Spodoptera frugiperda (Sf9) cells with wild-type Autographa californica nuclear polyhedrosis virus (AcNPV) DNA. The recombinant virus vAc373GM-CSF was identified and purified by several rounds of plaque hybridization. By assaying the culture medium, we demonstrated recombinant virus infected Sf9 cells expressing hGM-CSF. Recombinant hGM-CSFs with apparent molecular masses of 14.5, 15.5 and 16.5 kDa were detected by the Western blot method. All 3 forms have biological activity of hGM-CSF. Following N-glycanase treatment, a single band of 14.5–15.5 kDa appeared in SDS-PAGE. Western blot analysis of expression in Sf9 cells treated with tunicamycin revealed only the presence of the 14.5 kDa species. Thus, the signal sequence of recombinant hGM-CSF could be recognized and cleaved by infected insect cell and the resultant molecule secreted into the media.

【 授权许可】

Unknown