Activating transcription with light using caged DNA

Two significant challenges that must be overcome for successful in vivo gene therapy are the delivery of transgenes to a specific target cell population and the subsequent expression only within these cells. Non-specific delivery of “silent” genes followed by site-specific induction is one potential targeting strategy. In this report, the authors describe the inactivation and light-based re-activation of plasmid plasmids transcription using a photosensitive caging compound. Plasmids coding for green fluorescent protein (pGFP) were caged with 1-(4,5-dimethoxy-2-nitrophenyl)diazoethane (DMNPE). An in vitro transcription kit was used to generate mRNA from linearized native and caged pGFP. Reaction products were then analyzed by denaturing agarose gel electrophoresis. Caged plasmids did not produce mRNA in the in vitro reaction. However, exposure of caged plasmids to 365 nm light increased the amount of mRNA produced. These results suggest that addition of the DMNPE cage group to plasmid DNA is capable of blocking its transcription, and this blockage can be reversed with light exposure