Introduction: Regulative molecules like microRNAs play a very important role in gene regulation and cell development. In 2007, Yamanaka et al. reported the in vitro generation of stem-like cells by activating a set of specific genes such as Oct-4 and Sox-2. To systematically screen for effects of microRNA a screening system is required. Methods: We designed and created for each gene of interest a vector containing two luciferase-genes; one of them coupled with the 3´UTR sequence of each stem cell gene. Results: Using the FRT recombinase technology we were able to rapidly generate isogenic cell lines that behave like monoclonal cell lines. Conclusions: Combining the FRT and the dual luciferase technology allows rapidly generating a sensitive screening system for reliable capturing gene regulatory effects in an in vitro system ready to be screened by several hundred microRNAs.