伪狂犬病毒gD基因在转基因烟草中的表达

将猪伪狂犬病毒 (pseudorabiesvirus ,PRV)最主要的保护性抗原基因gD完整编码区亚克隆到修饰的植物双元表达载体pBI 35SL中 ,使其置于强启动子CaMV 35S doubleenhancer TEV 5′UTR下游 ,构建的转基因植物双元表达质粒经农杆菌介导转化烟草 .PCR检测叶片筛选阳性植株 ,Southern杂交进一步证实gD已整合到转基因烟草基因组中 .固相酶联斑点试验和Western印迹表明 ,gD在烟草获得正确表达并具有抗原性

Expression of the gD Gene of Pseudorabies Virus in Transgenic Tobacco

The expression of the immunogenic glycoprotein D(gD) of pseudorabies virus (PRV) in transgenic tobacco was reported. The modified binary vector pBI 35SL was constructed by replacing the single cauliflower mosaic virus (CaMV) 35 S and GUS gene of pBI121 with modified CaMV 35 S promoter from pRTL NS2. The resulting vector had a CaMV 35 S promoter with duplicated enhancer region fused to the tobacco etch virus (TEV) 5′ untranslated region (UTR). Plant expression vector pBI gD was made by sub cloning the 1 2 kb fragment coding the full length gD of PRV under the control of the modified CaMV 35 S promoter. Tobacco was transformed by co cultivating leaf discs with %Agrobacterium% strains harboring pBI gD . The regenerated transformants were analyzed by PCR and Southern hybridization. The results indicated that the gD gene was integrated into the genomic DNA of the transgenic tobacco. Dot ELISA and Western blotting further demonstrated the expression and antigenicity of gD in transgenic tobacco.