Construction and Expression in Pichia pastoris of an Eukaryotic Vector for Hepatitis B Virus X Gene

Objective:To construct and express a Pichia pastoris expression vector of HBV X protein for exploring the contribution of X protein to the chronic hepatitis and the hepatocellular carcinoma.Methods:X gene with XhoⅠ and XbaⅠ restriction enzyme sites was obtained by PCR, and subcloned into the pPICZαA vector.After identified by restrictive enzymes digestion and sequencing, reconstructed plasmid was transformed into P.pastoris GS115.Expression of X protein was assayed in P.pastoris GS115 culture supernatant by SDS and Western blotting.Results:The target gene X frag-ment about 465 bp was intergrated into the genome of P.pastoris GS115.The recombinant X protein with molecular weight of 17 kDa was secreted into the culture supernatant and was confirmed by Western blotting.Conclusion:P.pastoris expression vector pPICZαA-HBx was constructed successfully and recombinant X protein was expressed in P.pastoris GS115.