Abstract The placenta is a unique organ that develops during pregnancy and is essential for survival and growth of the developing fetus. A major obstacle in understanding the role of potential mediators of placental development has been the lack of a facile method for trophoblast gene manipulation in intact animals. Lentiviral infection of blastocysts in cell culture [ex vivo] and subsequent embryo transfer is an established technique that leads to trophoblast gene transfer. Despite its introduction several years ago, the use of this ex vivo embryo transfer approach to achieve trophoblast gene modulation has been limited. This is not likely due to a lack of need for these techniques, as it provides a powerful tool to assess placental development and dysfunction. The reasons for its limited use are likely due to the requirement for highly specialized staff, the technically involved process, and the need to culture blastocysts in vitro to infect before embryo transfer. To overcome the limitations of ex vivo gene transfer, we have devised and propose to validate a novel in utero trophoblast transgenesis approach, in which nonsurgical lentiviral infection of blastocysts occurs in utero and provides trophoblast-specific gene expression in naturally mated pregnant females. This method will provide the ability to evaluate numerous genes in a rapid and highly efficient way to study the role of the placenta in development and disease states. This method is straightforward, specific, does not require highly specialized staff, and eliminates the use of anesthetic. In addition, the rate of pregnancy and pups/pregnancy is the same naturally mated mice. Completion of the proposed aims will provide a new and powerful way to determine the processes that regulate placentation and underlying factors involved in the development of pregnancy-associated disorders in a highly accelerated manner.