TITLE:
Cloning, Expression, and Immunogenicity Analysis of the CpsE Protein from Group B Streptococcus Isolated from Tilapia (Oreochromis Niloticus)
AUTHORS:
Xi Fu, Kaiyu Wang, Jinlu Huang, Jun Wang, Hai Lian, Yang He, Lanmin Li, Kaiyu Wang
KEYWORDS:
Recombinant CpsE Expression; Purification; Polyclonal Antibody
JOURNAL NAME:
Engineering,
Vol.4 No.10B,
October
21,
2013
ABSTRACT: Group B Streptococcus (GBS) is a
major cause of serious bacterial infection in numerous animal species. The
production of capsular polysaccharide(CPs) is vital to GBS to evade host
immunity. One of the genes that required for production of CPs, cpsE, has been
determined to be well conserved in capsule gene cluster (cps).This study cloned
the cpsE gene from Tilapia of GBS clinical isolate (serotype Ia) and expressed
this gene with aid of pET-32a(+) in Escherichia coli BL21(DE3) competent cells
to obtain high levels of the recombinant protein for further study about CpsE
in fish and examination of its immunogenicity. The optimization of induction
conditions (IPTG concentration, temperature and time) in E.coli was accomplished
and let us to perform the recombinant protein induction at 37℃ for 3h,with 0.2mM IPTG in Luria Bertani (LB) medium.
At the optimal conditions, recombinant protein was expressed in an insoluble
form (inclusion bodies) and accounted for approximately 23% of the total
protein. Purification by affinity chromatography yielded about 480mg fusion
protein per liter culture.