Recent advances in Herpesvirus genetics using bacterial artificial chromosomes

Although the members of the Herpesvirus family are responsible for a wide v ariety of human diseases, advances in the understanding of viral molecular mechanisms of pathogenesis have been hampered by the large size of herpesvi rus genomes, rendering the viruses difficult to experimentally manipulate. Better techniques have been needed to facilitate mutagenesis of herpesvirus genomes, allowing for the assessment of the role of specific viral gene pr oducts in replication, immunity, and pathogenesis. Homologous recombination with plasmids containing genes of interest flanked by selectable markers h as been a successful method for generating viral mutants, as has the genera tion of recombinant virus from transfection of cosmid clones. Although thes e efforts to generate recombinant viruses have met with modest success, the protocols have been cumbersome. More recently, a novel technique for the m anipulation of herpesvirus genomes has been developed. This technology util izes bacterial F plasmids, and allows for the stable cloning of herpesvirus genomes as bacterial artificial chromosomes (BACs) in Escherichia coli. On ce cloned, such BACs are stable, and DNA purified from E. coli is infectiou s, fully capable of reproducing replication competent virus. Manipulation o f herpesvirus genomes is now feasible using the powerful techniques of bact erial genetics, and should facilitate a better understanding of the molecul ar pathogenesis of herpesvirus infections. (C) 2001 Academic Press.