2014•Xibei nongye xuebaoRequires access
Cloning of E2 Gene of Bovine Viral Diarrhoea Mucosal Disease Virus and Its Expression in Pichia pastoris
Yu‐Cheng Liu, Wang Guochao, Qingling Meng, Jun Qiao, Xuepeng Cai, HE Zhihao, Haibo Yang, Wan Xin Peng, Chuangfu Chen
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Abstract
According to BVDV-2 E2 gene sequences in GenBank,E2 gene of BVDV-2SW strain amplified using RT-PCR by specific primers.E2 gene was cloned into T vector and sequenced,and then subcloned into the yeast(P.pastoris)secretary expression vector pPIC9Kto generate the recombinant expression vector pPIC9K-E2.Recombinant plasmid pPIC9K-E2was liberalized by SalⅠdigestion and then electrically transformed into P.pastoris GS115.GS115-pPIC9K-E2recombinant yeast was screened by G418,and then induced with 1.5% methanol.The expression product was analyzed by SDS-PAGE and western blot.SDS-PAGE analysis confirmed that the expression of recombinant fusion protein has a mass molecule of 23.2ku;western blot showed that the recombinant protein can reacts with BVDV-2polyclonal antibody,confirming that the expression of recombinant E2protein has good reactogenicity.