Cloning and mutagenesis of a herpesvirus genome as an infectious bacterial artificial chromosome
M Messerle, I Crnkovic… - Proceedings of the …, 1997 - National Acad Sciences
M Messerle, I Crnkovic, W Hammerschmidt, H Ziegler, UH Koszinowski
Proceedings of the National Academy of Sciences, 1997•National Acad SciencesA 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 …
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 …
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.
National Acad Sciences