Gardella Gel Analysis to Detect Herpesvirus Saimiri Episomal DNA

Abstract

Herpesvirus saimiri (HVS) is capable of infecting a range of human cell types with high efficiency. The viral genome persists as high-copy-number, circular, nonintegrated episomes that segregate to progeny on cell division. This allows HVS-based vectors to transduce stably a dividing cell population and provide sustained transgene expression for an extended period of time both in vitro and in vivo. Moreover, the insertion of a bacterial artificial chromosome (BAC) cassette into the HVS genome simplifies the incorporation of large amounts of heterologous DNA for gene delivery. These properties offer characteristics similar to that of an artificial chromosome combined with an efficient delivery system. In this protocol, Gardella gel analysis is performed to determine whether the HVS genome is maintained in a nonintegrated episomal form. This method allows the identification of chromosomal/integrated DNA and episomal and linear forms of viral DNA, which run at the top, middle, and bottom of the gel, respectively. To obtain the required sensitivity to detect HVS episomes within host tissues, a modified polymerase chain reaction (PCR)-based Gardella gel can also be utilized.