Quantitative studies on the nuclear transport of plasmid DNA and gene expression employing nonviral vectors

The nuclear membrane is the main barrier to nonviral gene delivery. Thus, in the case of nondividing cells, a device for nuclear delivery of exogeneous DNA is necessary. In addition, to precisely evaluate the efficacy of various plasmid modifications and/or nonviral vectors, it is necessary to measure, not only gene expression but also the amount of delivered plasmid DNA into the subcellular compartment, particularly the nucleus. Moreover, it is also necessary to examine effects of the state of the plasmid DNA in the nucleus or various modifications of the plasmid DNA on the process after nuclear transport, i.e., transcription. Here, we address the issues of (1) the efficient delivery of genetic materials using a nuclear localization signal (NLS), (2) the quantitative evaluation of plasmid DNA delivered to the nucleus and the relationship between the amount of plasmid DNA delivered into the nucleus and gene expression, and (3) methods for evaluating of the effect of the state of plasmid DNA on transcription in vitro.