Expression of recombinant human zona pellucide 3 peptide in Pichia pastoris

Aim:In order to investigate the role of the human zona pellucide 3 portein(hZP3) in the fertility regulation,recombinant hZP3 C-terminal peptide was expressed in pichia pastoris yeast. Methods: The hZP3 DNA fragment(794-1282nt) was amplified by PCR from the hZP3 cDNA.The cloned DNA fragment of hZP3 was inserted into an expression vector pPICZα for pichia pastoris.Then,the linearized recombinant plasmid pPICZα-hZP3CT was transformed to Pichia pastoris X-33 by electroporation.The transformant cell carrying high copy hZP3CT gene were selected on the plates with high concentration zeocin.Expression of hZP3 peptide was induced by methanol and verified by SDS-PAGE and Western blotting.Results: The engineered yeast strain containing high copy-gene of the recombinant hZP3 peptide was successfully constructed.Western blotting showed the recombinant protein was expressed after methanol induction.Conclusion:The recombinant hZP3 peptide was successfully expressed in the engineered yeast and reacted specifically with the mouse antiserum against hZP3.