Expression of antimicrobial peptide Cecropin D in Pichia pastoris

To highly express antimicrobial peptide Cecropin D in Pichia pastoris,Cecropin D gene was amplified by SOEing PCR with 4 synthesized oligonucleotides and cloned into the pGAPZαA vector.The lineared pGAPZαA-CecropinD was transformed into P.pastoris SMD1168 by electroporation,the transformants were screened by Zeocin,it was proved that Cecropin D gene had intergrated into P.pastoris genome detected by PCR.Cecropin D had highly expressed under control of GAP promoter.The production could endure high temperature,acid and base,and it showed antibacterial activity to both Gram-positive and negative bacteria.The minimum inhibitory concentration(MIC) for E.coli DH5α and S.aureus Cowan I were 2.17 μg/mL and 4.55 μg/mL,respectively.