Cloning and expression of G2 glycoprotein gene of Hantavirus in Pichia pastoris

Objective: To study the cloning and expression of G2 glycoprotein gene of Hantavirus in methylotropic yeast Pichia Pastois. Methods: G2 glycoprotein gene was amplified by RT-PCR and tranfected into the competent E.Coli DH5α, which was confirmed by DNA sequence analysis.The G2 was cut by SnaBI and Not I and then ligated into pPIC9K phagemid vector which was digested with the same restriction enzymes. The expression vector pPIC9K-G2 was constructed and introduced into Pichia Pastois GS115 by electroporation after linearized with Sal I. The transformants were screened by G418, fermented in flasks and induced by 1% methanol. Results: The sequence analysis indicated that the sequence of cloned G2 gene was consistent with the design. The recombinant protein was 50KD by 12%SDS-PAGE.The high specificity by Western-Blot assay was proved. Conclusion: The G2 glycoprotein gene is successfully expressed in Pichia Pastoris GS115 expression system.