Gene therapy, the expression in cells of genetic material that has therapeutic activity, holds great promise for the treatment of a number of human diseases. A gene delivery vehicle, or vector, that may be of viral or non-viral origin, is generally used to carry the genetic material. Viral vectors have been developed that exclude immunogenic genes while taking advantage of the genes responsible for proficient integration of the viral genome into that of the host. In this way, viral vectors improve the probability of long-term expression of the therapeutic gene, whereas non-viral vectors, that are not as efficient at introducing and maintaining foreign gene expression, have the advantage of being non-pathogenic and non-immunogenic. Although thousands of patients have been involved in clinical trials for gene therapy, using hundreds of different protocols, true success has been limited. A major limitation of gene therapy approaches, especially when non-viral vectors are used, is the poor efficiency of DNA delivery to the nucleus; a crucial step to ensure ultimate expression of the therapeutic gene product. Here we review existing gene delivery approaches and, in particular, explore the possibility of enhancing non-viral gene delivery to the nucleus by incorporating specific nuclear targeting sequences in vectors, using a range of different strategies.