伪狂犬病病毒gE/TK基因缺失突变株的构建 |
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引用本文: | 田志军,仇华吉,倪健强,周艳君,王云峰,童光志.伪狂犬病病毒gE/TK基因缺失突变株的构建[J].中国预防兽医学报,2004,26(3):181-185. |
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作者姓名: | 田志军 仇华吉 倪健强 周艳君 王云峰 童光志 |
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作者单位: | 中国农业科学院,哈尔滨兽医研究所,兽医生物技术国家重点实验室,黑龙江,哈尔滨,150001 |
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基金项目: | 国家高技术研究发展计划(863计划) |
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摘 要: | 在伪狂犬病病毒转移载体pBdTK-Uni的多克隆位点中插入由SV40启动子控制下的IacZ基因表达盒,同时在右侧同源臂下游插入一个1.7kb的KpnI片段,构建成一个新的转移载体pUhi-LacZ.用该载体与Bartha-K61株基因组通过脂质体法共转染Vero细胞,经过10代蓝斑筛选纯化和PCR鉴定获得了一株稳定表达LacZ基因的伪狂犬病病毒gE/TK基因缺失突变株,命名为rPrV-LacZ.在不同的细胞(PK-15、IBRS-2、Vero和CEF)上,对该重组病毒与亲本病毒的增殖滴度和细胞病变进行比较,未见显著差异.结果表明转移载体pBdTK-Uni具有实用性,可用于构建伪狂犬病病毒基因工程活载体疫苗.
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关 键 词: | 伪狂犬病病毒 重组病毒 转移载体 LacZ基因 |
文章编号: | 1008-0589(2004)03-0181-05 |
修稿时间: | 2003年7月15日 |
Generation of a gE/TK-gene-deleted mutant derived from pseudorabies virus Bartha-K61 strain |
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Abstract: | A 1.7kb KpnI fragment and a LacZ gene expression cassette were inserted into the transfer vector pBdTK-Uni of pseudorabies virus (PrV), produced a transfer vector pUni-LacZ. The purified genomic DNA of Bartha-K61 strain and transfer vector pUni-LacZ were used to cotransfect Vero cells using lipofection transfection procedure. A recombinant virus stably expressing LacZ gene was generated after ten cycles of blue plague purification and PCR identification, and designated as rPrV-LacZ. Compared to its parental virus Bartha-K61 strain, rPrV-LacZ showed no obvious defferences in virus multiplication and cytopathogenic effect in defferent cell cultures (PK-15, IBRS-2,Vero and CEF). The results showed that the transfer vector pBdTK-Uni can be used for creation of PrV recombinants expressing foreign gene(s). |
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Keywords: | pseudorabies virus transfer vector recombinant virus LacZ gene |
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