Secretive expression of the fusion protein HSA-CNP in Pichia pastoris

Purpose To develop the recombinant expression of fusion protein human serum albumin(HSA)-type C natriutetic peptides(CNP)in Pichia pastoris.Methods The fusion gene of HSA(about 1 800 bp)-CNP(about 400 bp)was prepared with overlapping PCR to spang joint HSA genes and CNP genes.After digesting with EcoRⅠ and NotⅠ,the fusion gene was cloned into an expression vector pPIC9k.The plasmid of pPIC9K/HSA-GCSF was constructed and transformed into P.pastoris KM71 by electroporation.With the methanol solution as a inducer,the fusion protein was expressed from the recombinant P.pastoris KM71.Results The fusion gene of HSA-CNP was about 2 200 bp by electrophoresis and the nuclear acid sequence of the gene was correct.The fusion protein with about 80 000 that secreted in the recombinant P.pastoris KM71 was made of CNP and HSA by Western blot analysis or by the CNP radioimmunoassay.Conclusion The fusion protein HSA-CNP was successfully expressed in Pichia pastoris.