The fusion expression and purification of chicken β-defensin-1(Gal-1)gene in Escherichia coli

Endogenous antimicrobial peptide defensins are essential components of innate immunity.Defensins are distributed throughout the host to control invading micro-organisms.Chicken beta-defensin-1(Gallinacin-1,Gal-1) has antimicrobial activity against potentially pathogens and is worthwhile being investigated and developed.The mature fragment of Gal-1 gene was amplified from the plasmid of pGEM-T Easy-gal-1,and then inserted into the EcoRⅠ and BamHⅠ enzyme-cutting sites of pET-32a(+) plasmid.The recombinant vectors named pET-32a(+)-gal-1 were transformed into the competent cell E.coli BL-21.The positive clones were cultured and induced to express target protein by addition of(0.5 mmol) IPTG in LA.Expression products with molecular weight of approximately 25 kDa were examined by SDS-PAGE and Western Blot,which showed that gal-1 gene was expressed in the form of fusion protein.Densitometric scanning analysis demonstrated that the fusion protein accounted for about 15.5% of the total bacterial protein.The expressed fusion proteins were purified with 50% Ni-NTA resin affinity chromatography.Highly purified proteins were achieved in the eluate.Further study will be continued based on the results of this research.