Expression and identification of bovine tracheal antimicrobial peptide in Pichia pastoris

To express bovine tracheal antimicrobial peptide(bTAP) in methylotrophic yeast Pichia.pastoris,two pairs of primers were designed according to the published sequences for bTAP and the codon bias of P.pastoris.The bTAP gene was amplified by overlapping PCR and cloned into the vector pPIC9K to construct the yeast secretory expression vector pPIC9K-bTAP.The recombinant plasmid digested by restriction enzyme SalⅠ was transformed to P.pastoris GS115,and positive clones were screened with G418 resistance and was induced by 0.5% methanol for expression.SDS-PAGE confirmed the correct molecular weight of recombinant expression protein.Antibacterial assay showed that bTAP had strong antimicrobial activity against Staphylococcus aureus.