CpG-oligodeoxynucleotides (CpG-ODN) confer local antiviral properties when delivered to the genital mucosa of mice. CpG-ODN treatment also prevents the replication of the herpes simplex virus 2 (HSV-2) in the murine macrophage cell-line RAW 264.7. We hypothesized that this CpG-dependent antiviral system is the result of alterations in gene expression. In this study we used high density cDNA and oligonucleotide microarrays to survey the gene expression of the mouse genome over seven time points within a 24h period in order to elucidate a program of genes responsible for the antiviral effect. We used hierarchical and/or K-means clustering to reveal similarly regulated genes. Interferon-related genes including Stat-1 and -2, IRF-3 and -7, and several members of the powerful antiviral oligoadenylate synthetase (OAS) family were regulated by CpG-ODN. We also inhibited NFkappaB activation and showed that CpG-dependent cytokine secretion requires NFkappaB, but that the antiviral effect persists independently of NFkappaB activity.