Fusion expression of gallinacin-8 cDNA in Escherichia coli.

To evaluate the function and feasibility of manufacturing chicken β defensin by genetic engineering,the mature cDNA of chicken β defensins gal-8 was amplified from the plasmid pGEM-T Easy-gal-8,and then cloned into prokaryotic expression vector pGEX-6P-1.The recombinant vector pGEX-6P-1-gal-8 was transformed into the competent cell Escherichia coli BL21(DE3).The positive clones were cultured and induced to express target protein by IPTG.Fusion expression products were approximately 30ku in molecular weight,and examined by SDS-PAGE.Densitometric scanning analysis demonstrated that the fusion protein accounted for about 119mg/L of the total bacterial proteins.The expressed fusion protein was purified with immobilized glutathione affinity chromatography column,and a complete sophisticated gal-8 polypeptide was cut from fusion protein by the Prescission protease.Gal-8 mature peptide possessed the antimicrobial activity to Microccus flavus NCIB 8166 in the assay of drug susceptibility by agar diffusion method.