摘要
以伪狂犬病毒为载体表达其他病原体抗原蛋白是伪狂犬病疫苗研究的一个重要方向。根据已发表的伪狂犬病毒(PRV)Ra株TK基因序列设计2对引物,用PCR方法得到同源左右臂,克隆入PUC19载体中。利用平末端连接的方法将绿色荧光蛋白(EGFP)的基因表达盒插入到缺失位点,并在下游引入1个多克隆位点,构建缺失通用转移载体PUC19TK-EGFP。用脂质体转染试剂盒将PUC19TK-EGFP和PRV-Ra株的基因组共转染BHK细胞,以EGFP为标记基因,用噬斑法得到纯化的重组病毒,命名为TK/EGFP,为研制以伪狂犬病毒为载体的二价或多价基因工程疫苗奠定了物质基础。
Use of PRV as a system for expression of foreign antigens has been increasingly evaluated as an alternative to conventional vaccine production system. In this study,according to the Genbank open sequence,two pairs of primer were designed in order to obtain homology recombination arms of TK gene of PRV-Ra strain. Then both of them were inserted into PUC19 vector. A report gene expression cassette ,EGFP containing a multicloning sites,was inserted into PUC19HEK/TK to construct a universal transfer vector PUC19TK-EGFP. It was used to cotransfect BHK cell together with the genome ofPRV Ra strain ,a recombinat virus was selected and purified 3 times in BHK cell through EGFP gene and named TK/EGFP. The above results demonstrated that the universal transfer vector might be contributive to developed bivalence or multivalence genetically engineering vaccine in the future.
出处
《现代农业科技》
2009年第15期308-310,共3页
Modern Agricultural Science and Technology
