【摘要】 根据GenBank上WSSV囊膜蛋白基因vp19的序列,设计并合成引物,PCR扩增得到vp19基因并克隆到pGEM‐T载体中,经过BamHⅠ/HindⅢ酶切、连接并将vp19插入到pET32b表达载体中。用重组质粒pET32b-vp19转化大肠杆菌Origam(iDE3)pLysS,在IPTG诱导下,融合蛋白Trx-VP19以可溶性的形式得到表达,经SDS-PAGE和Western-blot检测显示其分子量与预期的大小相符合。目的蛋白经Ni2+柱纯化并定量后分别直接注射鳌虾和包被饲料投喂鳌虾。实验结果表明注射Trx-VP19可以提高鳌虾个体抗WSSV感染力的作用。更多还原

【Abstract】 The vp19 gene of White spot syndrome virus (WSSV) was amplified by PCR with specific primers and subcloned into the prokaryotic expression vector pET32b to get the recombinant plasmid pET32b-vp19. A positive plasmid was transformed into the host cell Origam(iDE3)pLysS and the target gene was successfully expressed as a fusion protoen when induced with IPTG. The molecular weight of the engineered protein was about 37kDa as demonstrated by SDS-PAGE and Western-blot. The engineered protein Trx-VP19 purified by Ni2+-column chromatography was injected into crayfish or used as a supplement to feed crayfish. Crayfish vaccinated by intramuscular injection with Trx-VP19 showed lower cumulative mortality due to WSSV compared to vaccination with bacteria expressing the empty vectors.更多还原