Cloning and mutagenesis of a herpesvirus genome as an infectious bacterial artificial chromosome
Cloning and mutagenesis of a herpesvirus genome as an infectious bacterial artificial chromosome
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DOI：
10.1073/pnas.94.26.14759
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发表时间：
1997-12-23
影响因子：
11.1
通讯作者：
Koszinowski, UH
中科院分区：
文献类型：
--
作者：
Messerle, M;Crnkovic, I;Koszinowski, UH
描述了一种感染性疱疹病毒基因组的克隆和诱变策略。将小鼠巨细胞病毒基因组克隆到大肠杆菌中，构建成230kb的细菌人工染色体（BAG）。杆菌将BAC质粒转染到真核细胞中导致生产性病毒感染。将靶向突变引入BAC克隆病毒基因组的可行性通过立即早期1基因的突变和突变病毒的产生来显示。因此，突变疱疹病毒基因组的完整构建现在可以在感染性子代重建之前以受控的方式进行。所描述的方法应该普遍适用于其他大DNA病毒基因组的诱变。
A strategy for cloning and mutagenesis of an infectious herpesvirus genome is described. The mouse cytomegalovirus genome was cloned and maintained as a 230 kb bacterial artificial chromosome (BAG) in E. coli. Transfection of the BAC plasmid into eukaryotic cells led to a productive virus infection. The feasibility to introduce targeted mutations into the BAC cloned virus genome was shown by mutation of the immediate-early 1 gene and generation of a mutant virus. Thus, the complete construction of a mutant herpesvirus genome can now be carried out in a controlled manner prior to the reconstitution of infectious progeny. The described approach should be generally applicable to the mutagenesis of genomes of other large DNA viruses.