In vivo electroporation is a powerful technique for the introduction of genes into organisms. Temporal and spatial regulation of expression of introduced genes, or of RNAi, would further enhance the utility of this method. Here we demonstrate conditional regulation of gene expression from electroporated plasmids in the postnatal rat retina and the embryonic mouse brain. For temporal regulation, Cre/loxP-mediated inducible expression vectors were used in combination with a vector expressing a conditionally active form of Cre recombinase, which is activated by 4-hydroxytamoxifen. Onset of gene expression was regulated by the timing of 4-hydroxytamoxifen administration. For spatial regulation, transgenes were expressed by using promoters specific for rod photoreceptors, bipolar cells, amacrine cells, Müller glia or progenitor cells. Combinations of these constructs will facilitate a variety of experiments, including cell-type-specific gene misexpression, conditional RNAi, and fate mapping of progenitor and precursor cells.