Generation of Transgenic Mice by Pronuclear Microinjection

Abstract

The introduction of a transgene into a fertilized oocyte by pronuclear microinjection of a solution containing the construct of choice is probably the most straightforward method to generate a genetically modified organism. This technique has been adapted to a number of vertebrate and invertebrate species and readily yields founder animals carrying the transgene if performed correctly. We will here describe the generation of transgenic mice from small transgenes and bacterial artificial chromosome (BAC) type transgenes by random integration. Moreover, we provide a method for the transposase-mediated integration of a transgene that is flanked by transposase recognition sequences. While not all species require the use of a sophisticated setup, the generation of transgenic mice is technically challenging mainly because of the small size of the oocyte and the need for well-defined buffer and media conditions. Moreover, manipulated embryos have to be put back into an environment where they can develop to term, and this environment can only be the oviduct of a recipient mouse that has been prepared to allow pregnancy.