黄颡鱼免疫球蛋白M重链基因原核表达研究

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Abstract

Yellow catfish (Pelteobagrus fulvidraco) is an important cultured catfish in China, and suffering from various diseases. Immunoglobulin M (IgM) is an important molecule against pathogens in fish immune response. In order to elucidate its expression pattern and mechanism during the period of immunological reaction for prevention of fish diseases, it is necessary to obtain enough IgM protein for preparation of anti-IgM specific antibodies. Therefore, the recombinant expression plasmid (IgM-pQE30) of yellow catfish was constructed with the IgM heavy chain coding sequence fragment cloned from the catfish linking the prokaryotic expression vector pQE30 after complete digestion of sequencing plasmid (IgM heavy chain/pMD-18T) and pQE30 by two endonucleases Sph 1 and Kpn 1. Then the recombinant plasmid was transferred into the host bacterium Escherichia coli M15, and induced to express fusion protein, which was confirmed by means of SDS-PAGE and Western blotting. A 62 kDa protein was greatly induced in unsoluble status in M15, and confirmed to be the recombinant protein (target protein) using monoclonal antibody against 6xHis tag in Western blotting. The recombinant protein was expressed in the greatest quantity at 30℃ for the induction of 8 hours with 0.2 mmol/L IPTG. The IgM heavy chain fusion protein can be greatly expressed in the recombinant plasmid expression system under the most suitable induction conditions mentioned above. This lays a foundation for further researches on the IgM of yellow catfish.

Key words： immunoglobulin M heavy chain recombinant plasmid prokaryotic expression Pelteobagrus fulvidraco

Received: 25 February 2013

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	YU Bo
	ZHANG Qi-zhong

Cite this article:

YU Bo,ZHANG Qi-zhong. Prokaryotic expression of immunoglobulin M heavy chain gene from yellow catfish(Pelteobagrus fulvidraco)[J]. , 2013, 32(1): 44-50.

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http://www.ecolsci.com/EN/ OR http://www.ecolsci.com/EN/Y2013/V32/I1/44

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