Aim: In order to express a novel gene named as BCL-G(L); of swine in E.coli and prepare its polyclonal antibody.
Methods: The contig sequence of the gene was predicted and in silicon cloned by blasting the human BCL-G(L); in swine ESTs database in NCBI. The cloning sequence was obtained by RT-PCR from swine spleen. The cloning sequence was identified by sequencing and compared with the contig sequence. Then the gene was cloned into a prokaryotic expression vector pET-32a to construct a recombinant plasmid named as pET32a-BCL-G(L);. The fusion protein pET32a-BCL-G(L); was expressed in E.coli BL21 and purified using a His-tag fusion protein purification kit. Then guinea pigs were immunized with the purified protein to get the specific polyclonal antibody.
Results: The titer of the antibody was 1:800 detected by ELISA. The protein BCL-G(L); can be specifically detected by western blot assay using the polyclonal antibody.
Conclusion: The novel swine gene BCL-G(L); was cloned and expressed in E.coli and its polyclonal antibody was prepared successfully.