Expression of Human Interleukin-13 In E. coli and Insect Cell Systems.

• Main Authors: Sheen, B. N., 沈柏年
• Other Authors: K.L.Lin, K.Y.Tray
• Format: Others
• Language: zh-TW
• Online Access: http://ndltd.ncl.edu.tw/handle/22582017855239934922

碩士 === 中山醫學院 === 生物化學研究所 === 84 === Interleukin 13 (IL-13)is a recently found cytokine secreted by activated T cells that is a potent in vitro modulator of human monocyteand B cell functions. The studies of IL-13 and IL-4 indicate that they share many common biological activites. Both IL-13 and IL-4 inhibit the production of proinflammatorycytokines, chemokines and hemotopoietic growth factorsby human monocyte, and can also enhance the Ig isotype class switching and the secretion of IgE and IgG4. These indicate that IL-13 play an important role in anti-inflammatory responses, allergic response and many clinical diseases. In order to further investigate the functions and roles of hIL-13, we hope to obtain a large quantify and reliable source of hIL-13. Firstly, we use a E. coli/pET-30 expression system to produce recombinant hIL-13 protein. hIL-13 DNA wsa ligated with pET-30 plasmid and transformed them into E. coli BL21 (DE3). hIL-13 fusion protein was purified using His-Binding Resin. Since hIL-13 is an eukaryotic protein, we also use a Baculovirus/insect cell system to express hIL-13. Two transfer plasmids (pVL1393, pAcG2T) was ligatedwith hIL-13 cDNA, respectively, and co-transfected the ligands with Baculovirus DNA into SF-9 inscet cell line. In this eukaryotic system, we wish to obtain recombinant hIL-13 that will resemble its native confer part. The biological activity of the recombinant hIL-13 was determined by using TF-1 cell line and human monocytes.