SUB-NUCLEAR VISUALIZATION OF RECOMBINATION BETWEEN TWO INCOMING HSV-1 GENOMES

Homologues recombination (HR) between incoming viral genomes is considered to be a major driving force for evolution and population diversification of DNA viruses. Despite the importance of HR, little is known about the nature physical interaction between incoming viral genomes. Herpes Simplex 1 (HSV-1) is a large double stranded DNA virus highly prevalent in human population and is a prototype virus for understanding herpesviridae replication. HSV-1 DNA replicates inside the nucleus as distinct foci called replication compartments (RCs) whereas each RC is originated from the genome of a single virion. This have led us to question, where and when during the replication process does the viral genomes come into contact to enable HR? We have developed a novel method based on Fluorescent In-Situ hybridization (FISH) to visually differentiate between incoming HSV genomes. In order to study recombination events, we have visualized co-infecting viral genomes at the sub-nuclear level. Analysis of the images generated suggests HR begins to occur at the early stages of DNA replication on the edges newly formed RCs. Variation in recombination frequency can be detected in different cell lines both by the FISH assay and by traditional population based assay. Our results indicate that the FISH assay can provide a new method to study recombination events in the single cell level.