关键词: 鸭, MHCⅡβ基因, 克隆, 原核表达, 多克隆抗体

Abstract: According to mRNA gene sequence registered in GenBank, a pair of specific primers for the gene of duck MHCⅡβ chain was designed and synthesized. Using total RNA from duck spleen, the target gene fragment was obtained by RT-PCR, then cloned into the T cloning vector pMD18-T. After being identified by restriction digestion and DNA sequencing, the insert fragment was subcloned into the expression vector pGEX-KG. After transforming into E.coli BL21, the recombinant plasmid were induced to obtain the interest protein. Then the purified protein was immunize to mouse for preparing polyclonal antibody. The results showed that MHCⅡβ gene was cloned, the obtained 798 bp fragment has 92% identities to the previously identified duck MHCⅡat nucleotide level; The prokaryotic expression vector was successfully constructed and the protein was expressed efficiently. The purity of protein reached 95%. A high titer and specific antibody has been prepared by purified protein, it was proved by the methods of ELISA and Western blot. All this makes it possible to do further studies on duck MHCⅡgene.

Key words: duck, MHCⅡβ gene, cloning, prokaryotic expression, polyclonal antibody