Abstract:
In order to obtain the proteins and polyclonal antibodies of the three subunits HblA,HblC and HblD of Bacillus cereus hemolysin BL,the PCR method was used in this experiment to amplify the HblA,HblC and HblD gene fragments for the construction of recombinant expression plasmids based on the prokaryotic expression vector pET-28 a. E. coli BL21 competent cells were used to express HblA,HblC and HblD respectively, and the prokaryotic expression proteins were purified by Ni
2+ agarose column; Balb/c mice were immunized with the purified proteins for the preparation of the corresponding polyclonal antibody; hemolysin BL in culture supernatant of Bacillus cereus BC12 was detected by western-blot. The results showed that the recombinant expression plasmids of HblA,HblC and HblD genes were successfully constructed; the recombinant proteins HblA,HblC and HblD of the same size as expected were obtained, which were 44.7,52.1,46.8 ku, respectively; polyclonal antibodies of HblC and HblD were prepared, and both HblC and HblD polyclonal antibodies could specifically recognize the BL subunit of hemolysin in the culture supernatant of Bacillus cereus. The results suggested that the polyclonal antibody prepared in the experiment could be used for the detection of Bacillus cereus hemolysin BL.