Co-expression of porcine reproductive and respiratory syndrome virus GP5 and M proteins in prokaryotic expression system

• Main Author: 洪嘉佑
• Other Authors: 吳美莉
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/09104961840301672805

碩士 === 國立屏東科技大學 === 生物科技研究所 === 100 === Porcine reproductive and repiratery syndrome virus (PRRSV), an aggressive virus can infects porcine alveolar macrophages. PRRS is the most serious disease that affects economically pig production in the world. PRRSV is an enveloped, positive-sense single-stranded RNA virus belonging to the Arteriviridae family. The genome of PRRSV is approximately 15 kb in length and contains nine open reading frames. The glycoprotein 5 (GP5) and non-glycosylated matrix protein (M) exist as a disulfide-linked heterodimer in the viral surface and has been indicated to elicit a protective antibody response to protect against PRRSV. However, the decoy epitope located at the N-terminus of GP5, might reduce the strength of the neutralizing antibody response against PRRSV. To develop a more efficient subunit vaccine against PRRS, double-gene plasmids which contained M and GP5 or dNGP5 (GP5 without decoy epitope) genes have been constructed and expressed in Escherichia coli system. In the study, M and GP5 or dNGP5 were cloned into pET21b vector under the control of two T7 promoters. SDS-PAGE and Western blot analysis were used to confirm the co-expression of M and GP5 or dNGP5. Furthermore, the genes fusions of GP5-M and dNGP5-M were also displayed on the cell surface of E. coli by using the N-terminal region of ice nucleation proteins. SDS-PAGE and Western blot analysis identified the synthesis of INPN-GP5-M fusion protein.