Isolation of Herpes Simplex Virus Nucleocapsid DNA

As an inanimate virus, herpes simplex virus type 1 (HSV-1) necessarily encodes all of its functions in its DNA. Isolation of pure viral DNA allows multiple downstream applications, including the creation of recombinant HSV strains, cloning of selected regions, and sequencing of viral DNA. The term nucleocapsid refers to the combination of the viral genome with the enclosing capsid; these viral genomes are necessarily linear and have been packaged for egress, even if they are not yet released from the cell. In contrast, viral DNA that is not associated with capsids may include episomal or concatenated forms and may have modifi-cations such as histones that are added within cells. During this protocol, the viral capsid protects the HSV genome from reagents that strip away and destroy most cellular contaminants. This procedure describes the isolation of viral nucleocapsids and their subsequent dissolution to purify clean, linear HSV DNA. Key words Nucleocapsid DNA, Genomes, Centrifugation, Nonionic detergents, Freon