ORCID
- Michael A. Jarvis: 0000-0002-0124-4061
Abstract
Herpesviruses are large DNA viruses and include important human and veterinary pathogens. Their genomes can be cloned as bacterial artificial chromosomes (BACs) and genetically engineered in Escherichia coli using BAC recombineering methods. While the recombineering methods are efficient, the initial BAC-cloning step remains laborious. To overcome this limitation, we have developed a simple, rapid, and efficient BAC-cloning method based on single-step transformation-associated recombination (STAR) in Saccharomyces cerevisiae. The linear viral genome is directly integrated into a vector comprising a yeast centromeric plasmid and a BAC replicon. Following transfer into E. coli, the viral genome can be modified using standard BAC recombineering techniques. We demonstrate the speed, fidelity, and broad applicability of STAR by cloning two strains of both rat cytomegalovirus (a betaherpesvirus) and Kaposi's sarcoma-associated herpesvirus (a gammaherpesvirus). STAR cloning facilitates the functional genetic analysis of herpesviruses and other large DNA viruses and their use as vaccines and therapeutic vectors.
DOI
10.1016/j.crmeth.2024.100696
Publication Date
2024-02-26
Publication Title
Cell Reports Methods
Volume
4
Issue
2
Keywords
BAC, CP: Microbiology, DNA virus, herpesvirus, HHV-8, KSHV, RCMV, recombination, YAC, YCp, yeast
Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.
Recommended Citation
Brune, W., Knickmann, J., Staliunaite, L., Puhach, O., Ostermann, E., Günther, T., Nichols, J., Jarvis, M., Voigt, S., Grundhoff, A., & Davison, A. (2024) 'A simple method for rapid cloning of complete herpesvirus genomes', Cell Reports Methods, 4(2). Available at: https://doi.org/10.1016/j.crmeth.2024.100696
