Mouse B7H4 prokaryotic expression vector expression and polyclonal antibody preparation.doc

 PAGE \* MERGEFORMAT 15 Mouse B7H4 prokaryotic expression vector expression and polyclonal antibody preparation Of: Hayshed dry, Xiao Huan, Hu Guoyan, that Zhiping, Zheng Shuhua, Liu Wei, Zhang Liangqing, Xu Jun-Fa [Abstract] Objective To investigate the expression of mouse B7 H4 prokaryotic vector, expression and polyclonal antibody preparation. Methods specific primer mouse B7 H4 (mB7 H4 functional areas of extracellular DNA, by enzyme digestion, splicing prokaryotic expression vector pET28a mB7 H4. The recombinant plasmid was transformed into E.coli BL21 (DE3) strain induced by IPTG, Ni NTA affinity chromatography purified proteins, SDS PAGE analysis of protein purity. The purified protein immune rabbit polyclonal antibody, and its purification and identification. The results of sequencing confirmed that the construction of recombinant expression vector pET28a mB7 H4 mB7 H4 coding sequence with its sequence comparison with GenBank published sequences of the same plasmid in E. coli induced to express the relative molecular mass (Mr) for the purpose of 26.5 KD protein, SDS PAGE analysis showed that the purified protein to electrophoretic homogeneity. double agar diffusion test titer of 1:16, ELISA titer assay 1:12 800, Western blot analysis showed that antibodies can specifically bind to mB7 H4. Conclusions The high expression of recombinant prepared mB7 H4 protein in prokaryotic expression vector and high titer of anti mB7 H4 polyclonal antibody for further study of the function of B7 H4 laid basis. [Keywords:] B7 H4, prokaryotic expression vector, gene expression, polyclonal antibody Abstract: Objective To study the construction and expression of a prokaryotic expression vector for murine B7 H4 and preparation of a polyclonal antibody.Methods Murine B7 H4 (mB7 H4) gene was amplified with specific primers. The PCR product was digested with enzymes and then ligated to reconstruct recombinant prokaryotic expression vector pET28a mB7 H4 that expresses