Messerle M, Crnkovic I, Hammerschmidt W, Ziegler H, Koszinowski U H
Max von Pettenkofer-Institut für Hygiene und Mikrobiologie, Ludwig-Maximilians-Universität München, Feodor-Lynen-Strasse 25, D-81377 Munich, Germany.
Proc Natl Acad Sci U S A. 1997 Dec 23;94(26):14759-63. doi: 10.1073/pnas.94.26.14759.
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 (BAC) 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.
描述了一种用于感染性疱疹病毒基因组克隆和诱变的策略。小鼠巨细胞病毒基因组被克隆并作为一个230 kb的细菌人工染色体（BAC）保存在大肠杆菌中。将BAC质粒转染到真核细胞中可导致产生有活性的病毒感染。通过立即早期1基因的突变和突变病毒的产生，证明了将靶向突变引入BAC克隆病毒基因组的可行性。因此，现在可以在有感染性后代重建之前，以可控的方式完成突变疱疹病毒基因组的完整构建。所描述的方法应该普遍适用于其他大型DNA病毒基因组的诱变。
