Prokaryotic Expression of Qiongzhuea tumidinoda CBF1 Gene and Preparation of Its Polyclonal Antibody

C-repeat binding factor 1 gene(CBF1)was amplified by RT-PCR from Qiongzhuea tumidinoda Hsueh leaves and cloned into prokaryotic expression vector pET32a(+). After identification by enzyme digestion and sequencing,the expression of recombinant plasmid carried CBF1 gene was transformed into Rosetta2(DE3)E. coli. Through induced with IPTF,the expression of recombinant protein was analyzed by SDS-PAGE. The results showed that the protein was highly expression in E. coli, and the molecular weight of the recombinant protein was 45 kD. After purification with Ni2+-NTA affinity chromatography,the immune reactivity of recombinant CBF1 protein was identified with positive antiserum against nature CBF1 protein specifically by Western-blotting analysis. Antibodies against recombinant CBF1 protein was obtained by subcutaneous injection of rabbit with purified CBF1 recombinant protein,which is specific to the total protein of Qiongzhuea tumidinoda Hsueh leaves induced by low temperature. Our results indicate that the recombinant protein can be used for immunohistochemistry and western blot detection.