Objective To clone, express and purify C12orf49 recombinant protein. To prepare rabbit anti-C12orf49 protein polyclonal antibody in order to further elucidate its biological function.
Methods PCR was used to amplify the gene C12orf49 in vitro. pET-32a (+)-C12orf49, the recombinant protein prokaryotic expression vector, was transformed into E. coli. IPTG was used as the inductive agent to obtain C12orf49 recombinant protein, and the recombinant protein was analyzed with sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE) and Western blot. Specific polyclonal antibody was derived from rabbits that immunized by recombinant protein. ELISA and Western blot were used to test its titer and specificity, respectively. MTT cell proliferation experiment was carried out to observe effect of the protein on proliferation of HepG2 cells.
Results The C12orf49 recombinant protein was expressed in a large quantity. Data of ELISA indicated that the titer of polyclonal antibody was higher than 1:1 280 000. And the antibody also had a good specificity, confirmed by Western blot. C12orf49 recombinant protein may had a advanced effect on the proliferation of HepG2 cells.
Conclusions Using C12orf49 recombinant protein, we can obtain the polyclonal antibody with great titer and good specificity. Human novel gene C12orf49 encoded protein could promote the proliferation of HepG2 cells.