关键词: 小鼠BPI_36-259目的蛋白, E.coli BL21(DE3), 多克隆抗血清

Abstract: Objective To make murine BPI_36-259 protein with prokaryotic expressing system and prepare rabbit polyclonal antiserum with the recombinant protein. Methods Functional domain of murine BPI was predicted by bioinformatics methods. cDNA of muBPI_1-281 was synthesized and cloned to pUC57 cloning vector to generate pUC57-muBPI_1-281 construct. Then muBPI_36-259 functional fragment was amplified on the pUC57-muBPI_1-281 plasmid by PCR and cloned into prokaryotic expression vector, pET28a, to produce the expression vector pET28a-muBPI_36-259. After IPTG inducing, recombinant protein was expressed in bacteria E.coli BL21(DE3) in the main form of inclusion bodies. This recombinant protein was used to immunize rabbit to generate polyclone anti-serum, the titer and the specificity of antiserum was detected by indirect ELISA method. Results Prokaryotic expression construct at murine BPI functional domain(pET28a-muBPI_36-259) has been successfully generated, which express this recombinant protein in the main form of inclusion bodies. Using this recombinant protein to immunize rabbit, we got specific anti-muBPI_36-259 antiserum with the titer of 1∶10 000. Conclusion We have obtained the antibiosis protein(muBPI_36-259) in main form of inclusion bodies from prokaryotic expression system and used it to immunize rabbit, the specific antiserum can be used to detect the target antibiosis protein of muBPI_36-259.

Key words: muBPI_36-259 recombinant protein, E. coli BL21(DE3), polyclonal antiserum