EXPRSSION OF HUMAN ENDOSTATIN IN PICHIA PASTORIS

Objective:To gain recombinant endostatin in Pichia pastoris.Methods:The total mRNA was isolated from Chinese fetal liver,endostatin cDNA was synthesized with RT PCR,then cloned into pGEM T vector.The positive clones were screened with PCR,restriction map and DNA sequencing.Endostatin cDNA was subcloned into Pichia expression vector pGAPZa.Pichia pastoris GS115 cells were transformed with recombinant DNA.SDS-PAGE was used to assay the expression of endostatin in recombinant GS115.After purified with G25 and heparin,the recombined protein was used in the proliferation assay of ECV304 cells.Results:DNA sequencing result showed the endostatin cDNA was successfully cloned.After transformation with recombined DNA,GS115 cells expressed recombinant human endostatin.MTT assay implicated the recombinant protein could inhibited the proliferation of ECV304 cell sejectivity.Conclusion:Recombinant human endostatin protein was sa cesefuely expressed in Pichia pastoris.