关键词: 鸡, 骨保护素, 原核表达, 纯化, 抗体

Abstract: Osteoprotegerin(OPG) is a new found protein which can inhibit the osteoclast differentiation and activation. In order to do further research about its biological function, the total RNA were extracted from osteoblasts cultured in vitro for OPG amplification by RT-PCR method. After the sequence testing was correct, the OPG gene was inserted to the expression plasmid pET-32a(+) and expressed in Rosetta-gami（DE3）pLysS with IPTG inducement. The SDS-PAGE result showed that the cloned recombinant protein expressed in the form of inclusion bodies in Rossetta with molecular weight of 63kD and amounted to 11.9% of the whole protein, and western blotting indicated that the expressed protein had satisfied immunobiological activity. Pure protein was obtained by Ni2+-NTA chelating column for preparation of anti-OPG polyclonal antibody by immunizing rabbit, the titer of antiserum generated was 1:10240 by ELISA. Western blotting analysis showed that it could bind with OPG protein specially expressed in Pichia Yeast.

Key words: Chicken, OPG, Prokaryotic Expression, Purification, Antibody