Stable Insertion into the Human Cytomegalovirus Genome of a Foreign Gene.

外源基因稳定插入人类巨细胞病毒基因组。

• 批准号: 01480182
• 负责人: WATANABE Yasushi
• 金额: $ 4.29万
• 依托单位: Tokai University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-01480182/
• 关键词: Human cytomegalovirus; Expression vector; Recombinant virus; Vaccine

On the bases of previous finding that the HindIII O fragment of 7.8-kb in the genome of human cytomegalovirus strain Towne is nonessential for viral replication, we constructed a vector pKM that directs introduction of foreign genes by homologous recombination precisely replacing the O fragment. Using the vector, we constructed strain Towne-derived recombinant viruses in which a chimeric lacZ gene fused to the SV40 promoter and poly A signals was inserted in place of the O fragment. Two types of recombinants were obtained which carry the chimeric gene in opposite directions. Beta Gal was expressed in HEL cells infected with these recombinants throughout the infection cycle, and the rate of its synthesis in early stages of infection was comparable to that of synthesis of a 65 kDa viral glycoprotein, one of viral proteins expressed abundantly. The chimeric lacZ gene introduced was stable and no lacZ- revertant has been observed so far.

基于先前发现人巨细胞病毒Towne株基因组中7.8 kb的HindIII O片段对于病毒复制不是必需的，我们构建了精确替换O片段的通过同源重组引导外源基因导入的载体pKM。使用该载体，我们构建了源自 Towne 的重组病毒株，其中插入了与 SV40 启动子融合的嵌合 lacZ 基因，并插入了多聚 A 信号来代替 O 片段。获得了两种类型的重组体，它们携带相反方向的嵌合基因。 Beta Gal 在整个感染周期中在用这些重组体感染的 HEL 细胞中表达，其在感染早期的合成速率与 65 kDa 病毒糖蛋白（一种表达丰富的病毒蛋白）的合成速率相当。引入的嵌合lacZ基因是稳定的，迄今为止尚未观察到lacZ回复体。

项目成果

Masataka Takekoshi: "Site-specific stable insertion into the human cytomegalovirus genome of a foreign gene under the control of the SV40 promoter" Gene. (1991)

Masataka Takekoshi：“在 SV40 启动子控制下，外源基因在人类巨细胞病毒基因组中进行位点特异性稳定插入”基因。

竹腰 正隆: "EB・サイトメガロウイルス" 臨床と微生物. 17. 45-50 (1990)

Masataka Takekoshi：“EB/巨细胞病毒”临床微生物学 17. 45-50 (1990)。

Masataka Takekoshi: "Siteーspecific stable insertion into the human cytomegalovirus genome of a foreign gene under control of the Sv_<40> premoter" gene. (1991)

Masataka Takekoshi：“在 Sv_<40> 前驱基因的控制下，外源基因在人类巨细胞病毒基因组中进行位点特异性稳定插入”（1991）。

竹腰正隆: "EB・サイトメガロウイルス" 臨床と微生物. 17. 45-50 (1990)

Masataka Takekoshi：“EB/巨细胞病毒”临床和微生物学 17. 45-50 (1990)。

Masataka Takekoshi: "Siteーspecific stable insertion into the human cytomegalovirus genome of a foreign gene under control of the SV40 promoter" gene. (1991)

Masataka Takekoshi：“在 SV40 启动子的控制下，外源基因在人类巨细胞病毒基因组中进行位点特异性稳定插入”（1991）。