Eukaryotic expression of hepatitis E virus structural gene

Objective To express HEV structural gene by using pichia pastoris expression system. Methods The chimeric gene segment HEV ORF 23 (an 800bp gene segement close to the HEV ORF2 C end and the whole ORF3 gene segement) of HEV structural gene is cloned into pPICZ α A pichia expression vector by double digest with Not Ⅰ and Xba Ⅰ. The recombinant plasmid pPICZ α A_HEV ORF 23 is transformed appropriate pichia pastoris strain by using electroporation and then the interest gene is integrated into the genome DNA by a single crossover. The recombinant pichia strains are selected for induced expression by methanol, purifying the expression products and doing western_blot detection. Results We obtain a 69kDa protein in the cell lysates after induced 72 hours under pH6.0 circumstance. The protein can be purified with His·Bind Resin, and the western_blot results is positive. Conclusions Using pichia pastoris expression system and His purification system, a soluble, high antigenic and immunogenic HEV recombinant protein can be obtained, and it makes a foundation for the development of the efficient hepatitis E diagnostic reagent and vaccine.