Abstract: A UL9 substrate for detecting helicase activity in a UL9 protein is provided. In one embodiment, the substrate includes a first strand including a herpes replication origin sequence and a first single stranded tail; and a second strand including a sequence complementary to the herpes replication origin sequence. In one variation, the first single stranded tail is 3' relative to the herpes replication origin sequence. The second strand may optionally further include a single stranded tail. The single stranded tail on the second strand may be 3' or 5' relative to the sequence complementary to the herpes replication origin sequence. In another variation, the first single stranded tail is 5' relative to the herpes replication origin sequence. The second strand may optionally further include a single stranded tail. The single stranded tail on the second strand may be 3' or 5' relative to the sequence complementary to the herpes replication origin sequence.